Publications - Peer-reviewed articles

A randomized controlled trial investigating the neurocognitive effects of Lacprodan® PL-20, a phospholipid-rich milk protein concentrate, in elderly participants with age-associated memory impairment

Scholey, A. B., Camfield, D. A., Hughes, M. E., et al. — Tue, 26 Nov 2013 20:36:22 +0100

Background: Age-related cognitive decline (ARCD) is of major societal concern in an ageing population, with the development of dietary supplements providing a promising avenue for amelioration of associated deficits. Despite initial interest in the use of phospholipids (PLs) for ARCD, in recent years there has been a hiatus in such research. Because of safety concerns regarding PLs derived from bovine cortex, and the equivocal efficacy of soybean-derived PLs, there is an important need for the development of new PL alternatives. Phospholipids derived from milk proteins represent one potential candidate treatment.Methods: In order to reduce the effects of age-associated memory impairment (AAMI) the Phospholipid Intervention for Cognitive Ageing Reversal (PLICAR) was developed to test the efficacy of a milk protein concentrate rich in natural, non-synthetic milk phospholipids (Lacprodan® PL-20). PLICAR is a randomized, double-blind, placebo-controlled parallel-groups study where 150 (N = 50/group) AAMI participants aged > 55 years will be randomized to receive a daily supplement of Lacprodan® PL-20 or one of two placebos (phospholipid-free milk protein concentrate or inert rice starch) over a 6-month (180-day) period. Participants will undergo testing at baseline, 90 days and 180 days. The primary outcome is a composite memory score from the Rey Auditory Verbal Learning Test. Secondary outcomes include cognitive (verbal learning, working memory, prospective and retrospective memory, processing speed and attention), mood (depression, anxiety, stress and visual analogue scales), cardiovascular (blood pressure, blood velocity and pulse wave pressure), gastrointestinal microbiota and biochemical measures (oxidative stress, inflammation, B vitamins and Homocysteine, glucoregulation and serum choline). Allelic differences in the Apolipoprotein E and (APOE) and Methylenetetrahydrofolate reductase (MTHFR) gene will be included for subgroup analysis. A subset (N = 60; 20/group)) will undergo neuroimaging using functional magnetic resonance imaging (fMRI) and magnetoencephalography (MEG) in order to further explore in vivo central mechanisms of action of Lacprodan® PL-20. This study will enable evaluation of the efficacy of milk-derived phospholipids for AAMI, and their mechanisms of action.Trial Registration: The trial is jointly funded by Arla Foods and Swinburne University of Technology, currently recruiting and is registered on the Australian New Zealand Clinical Trials Registry as ACTRN12613000347763.

Dietary milk-fat-globule membrane affects resistance to diarrheagenic escherichia coli in healthy adults in a randomized, placebo-controlled, double-blind study

Ten Bruggencate, S. J., Frederiksen, P. D., Pedersen, S. M., et al. — Fri, 01 Jan 2016 20:36:22 +0100

Background: The milk-fat-globule membrane (MFGM) contains phospholipids and membrane glycoproteins that have been shown to affect pathogen colonization and gut barrier integrity. Objective: In the present study, we determined whether commercial heat-treated MFGM can increase resistance to diarrheagenic Escherichia coli. Methods: A randomized, placebo-controlled, double-blind, 4-wk parallel-intervention study was conducted in healthy adults. Participants were randomly assigned to a milk protein concentrate rich in MFGM [10 g Lacprodan PL-20 (Arla Foods Ingredients Group P/S), twice daily; n = 30; MFGM group) or a control [10 g Miprodan 30 (sodium caseinate), twice daily; n = 28]. After 2 wk, participants were orally challenged with live, attenuated diarrheagenic E. coli (1010 colonyforming units). Primary outcomes were infection-induced diarrhea and fecal diarrheagenic E. coli excretion. Secondary outcomes were gastrointestinal symptoms [Gastrointestinal Symptom Rating Scale (GSRS)], stool frequency, and stool consistency (Bristol Stool Scale). Results: Diarrheagenic E. coli resulted in increased fecal output, lower relative fecal dry weight, increased fecal E. coli numbers, and an increase in stool frequency and gastrointestinal complaints at day 1 after challenge. MFGM significantly decreased the E. coli-induced changes in reported stool frequency (1.1 ± 0.1 stools/d in the MFGM group; 1.6 ± 0.2 stools/d in the control group; P = 0.04) and gastrointestinal complaints at day 2 (1.1 ± 0.5 and 2.5 ± 0.6 GSRS scores in the MFGM and control groups, respectively; P = 0.05). MFGM did not affect fecal wet weight and E. coli excretion at day 2 after challenge. Conclusions: The attenuated diarrheagenic E. coli strain transiently induced mild symptoms of a food-borne infection, with complete recovery of reported clinical symptoms within 2 d. The present diarrheagenic E. coli challenge trial conducted in healthy adults indicates that a milk concentrate rich in natural, bioactive phospho- and sphingolipids from the MFGM may improve in vivo resistance to diarrheagenic E. coli. This trial was registered at clinicaltrials.gov as NCT01800396.

Effects of milk-based phospholipids on cognitive performance and subjective responses to psychosocial stress

Boyle, N. B., Dye, L., Arkbåge, K., et al. — Tue, 01 Jan 2019 20:36:22 +0100

Objectives: The aim of this study was to examine the stress-buffering potential of phospholipid (PL) intake on cognitive performance and neuroendocrine and psychological responses under conditions of psychosocial stress in a high-stress vulnerable (perfectionist) sample. Methods: Fifty-four high-perfectionist men consumed a 6-wk daily intake of a bovine milk–derived PL (2.7 g/d) or placebo drink in a randomized, double-blind, placebo-controlled, parallel groups design. Working memory, executive control function, and acute physiological/subjective responses to an acute psychosocial stressor were examined before and after the 6-wk PL or placebo intake. Results: PL intake improved post-stress reaction time performance on an attention-switching task (P = 0.01). No significant attenuation of the salivary cortisol stress response was shown. PL intake significantly increased mid-stress induction energetic arousal (P = 0.03). A non-significant reduction in anticipatory subjective stress was reported after PL intake (P = 0.06). Systolic and diastolic blood pressures (P < 0.04 and P = 0.01, respectively) were significantly augmented in the PL condition. Conclusions: Dietary intake of bovine milk PLs conferred cognitive performance benefit under conditions of psychosocial stress but failed to moderate cortisol response. Moderation of subjective response to stress exposure may have underpinned this performance protection.

Inorganic phosphate transporter PiT2 and phosphate homeostasis

Segelund, E. B., Kristensen, M. D., Holste, S., Lundby, J. M. B., Pedersen, L. — Thu, 07 Nov 2024 20:36:22 +0100

Exploiting O-GlcNAc dyshomeostasis to screen O-GlcNAc transferase intellectual disability variants

Yuan, H., Mitchell, C. W., Ferenbach, A. T., et al. — Tue, 14 Jan 2025 20:36:22 +0100

O-GlcNAcylation is an essential protein modification catalyzed by O-GlcNAc transferase (OGT). Missense variants in OGT are linked to a novel intellectual disability syndrome known as OGT congenital disorder of glycosylation (OGT-CDG). The mechanisms by which OGT missense variants lead to this heterogeneous syndrome are not understood, and no unified method exists for dissecting pathogenic from non-pathogenic variants. Here, we develop a double-fluorescence strategy in mouse embryonic stem cells to measure disruption of O-GlcNAc homeostasis by quantifying the effects of variants on endogenous OGT expression. OGT-CDG variants generally elicited a lower feedback response than wild-type and Genome Aggregation Database (gnomAD) OGT variants. This approach was then used to dissect new putative OGT-CDG variants from pathogenic background variants in other disease-associated genes. Our work enables the prediction of pathogenicity for rapidly emerging de novo OGT-CDG variants and points to reduced disruption of O-GlcNAc homeostasis as a common mechanism underpinning OGT-CDG.

Palatinale rugae til human identifikation:

Kofod Petersen, A., Bindslev, D. A., Villesen, P., Staun Larsen, L. — Fri, 08 Nov 2024 20:36:22 +0100

Background
The human palatal rugae (folds) comprise a detailed pattern in the hard palate. They are described to be very constant during an individual’s lifetime, comparable with a fingerprint, and, importantly within a forensic setting, can serve as an important supplement in odontological identification of deceased.
Aim
The aim of this study was to test two methods of superimposition in correctly distinguishing between matches (from same individual) and mismatches (from different individuals) of digital scans of rugae.
Methods
We used an existing research database holding digital intraoral scans of 51 individuals from two points in time approximately 6 months apart. The palatal rugae area was cut manually from the digital scans without regards to teeth position. Two superimposition methods (Iterative Closest Point (ICP) and Random Sample Consensus (RANSAC)) were tested in an all-vs-all manner. Similarity was reported as fitness and inlier Root Mean Squared Error (RMSE).
Results
For both ICP and RANSAC none of the similarity measures were able to unambiguously distinguish between matches (rugae from same individual at two time points) and mismatches (rugae from different individuals).
Conclusion
The results imply that the two superimposition methods are not applicable for matching rugae scans. Further exploration of possible methods to distinguish between matches and mismatches are needed to fully exploit the great potential within forensic odontology identification of these digital palatal 3D ‘fingerprints’.

Structure and Function of the Leukocyte Integrin αMβ2

Andersen, G. R., Emsley, J. — Sun, 01 Jan 2023 20:36:22 +0100

The integrin αMβ2 (also known as CD11b/CD18, Mac-1, complement receptor 3) is expressed on the surface of leukocytes and mediates numerous responses of these cells critical to innate immunity. The αMβ2 receptor contributes to the recruitment, firm adhesion, and transendothelial migration of leukocytes at sites of vascular injury and facilitates tissue inflammation. Biochemical and cell-based studies have characterized the interactions of the αMβ2 integrin with diverse ligands including plasma protein fibrinogen, complement protein fragment iC3b, and the cell surface receptors platelet glycoprotein Ib (GPIb) and intercellular adhesion molecule 1 (ICAM-1). The αMβ2 integrin exists in an inactive conformation and when activated by a variety of stimulae undergoes a structural change to an active form capable of binding to ligands with high affinity. Concurrently, allosteric changes occur upon ligand binding that result in “outside-in” cell signaling. Here we describe the αMβ2 protein structures and biophysical measurements that underpin the current understanding of diverse ligand recognition through the metal ion-dependent adhesion site (MIDAS).

Characterization of the bispecific VHH antibody tarperprumig (ALXN1820) specific for properdin and designed for low-volume administration

Tamburini, P., Pedersen, D. V., Devore, D., et al. — Mon, 01 Jan 2024 20:36:22 +0100

The bispecific antibody tarperprumig (ALXN1820) was developed as a treatment option for diseases involving dysregulated complement alternative pathway (AP) activity that could be administered in small volumes, either subcutaneously or intravenously. Tarperprumig incorporates a C-terminal variable domain of a heavy chain only antibody (VHH) that binds properdin (FP) connected via a flexible linker to an N-terminal VHH that binds human serum albumin (HSA). The purified bispecific VHH antibody exhibits an experimental molecular weight average of 27.4 kDa and can be formulated at > 100 mg/mL. Tarperprumig binds tightly to FP and HSA with sub-nanomolar affinity at pH 7.4 and can associate simultaneously with FP and HSA to form a ternary complex. Tarperprumig potently and dose-dependently inhibits to completion in vitro AP-dependent complement C5b-9 formation, AP-dependent hemolysis, and the AP deposition of C3, FP and C9. X-ray crystallography revealed that the isolated FP-binding VHH recognizes the thrombospondin repeat 5 domain of FP, thereby preventing FP from binding to the AP convertase owing to severe steric hindrance. Tarperprumig cross-reacts with cynomolgus monkey FP and serum albumin. In summary, tarperprumig exhibits properties tailored for subcutaneous administration and is currently in clinical development for the treatment of complement AP-related disorders.

Unveiling the enzymatic pathway of UMG-SP2 urethanase

Paiva, P., Teixeira, L. M. C., Wei, R., et al. — Wed, 01 Jan 2025 20:36:22 +0100

The recently discovered metagenomic urethanases UMG-SP1, UMG-SP2, and UMG-SP3 have emerged as promising tools to establish a bio-based recycling approach for polyurethane (PU) waste. These enzymes are capable of hydrolyzing urethane bonds in low molecular weight dicarbamates as well as in thermoplastic PU and the amide bond in polyamide employing a Ser-Ser cis -Lys triad for catalysis, similar to members of the amidase signature protein superfamily. Understanding the catalytic mechanism of these urethanases is crucial for enhancing their enzymatic activity and improving PU bio-recycling processes. In this study, we employed hybrid quantum mechanics/molecular mechanics methods to delve into the catalytic machinery of the UMG-SP2 urethanase in breaking down a model PU substrate. Our results indicate that the reaction proceeds in two stages: STAGE 1 - acylation, in which the enzyme becomes covalently bound to the PU substrate, releasing an alcohol-leaving group; STAGE 2 - deacylation, in which a catalytic water hydrolyzes the enzyme:ligand covalent adduct, releasing the product in the form of a highly unstable carbamic acid, expected to rapidly decompose into an amine and carbon dioxide. We found that STAGE 1 comprises the rate-limiting step of the overall reaction, consisting of the cleavage of the substrate's urethane bond by its ester moiety and the release of the alcohol-leaving group (overall Gibbs activation energy of 20.8 kcal mol -1). Lastly, we identified point mutations that are expected to enhance the enzyme's turnover for the hydrolysis of urethane bonds by stabilizing the macrodipole of the rate-limiting transition state. These findings expand our current knowledge of urethanases and homolog enzymes from the amidase signature superfamily, paving the way for future research on improving the enzymatic depolymerization of PU plastic materials.

Control of neonatal diarrhea in piglets with reduced antibiotic use by application of a complementary feed - a randomized controlled farm trial

Sall, K. K., Foldager, L., Delf, C., et al. — Fri, 10 Jan 2025 20:36:22 +0100

The objective of this farm trial was to investigate if the consumption of antibiotics could be reduced when piglets showing early signs of neonatal diarrhea were treated with an oral dose of tannin extract derived from sweet chestnut wood. The farm had a very high incidence of neonatal diarrhea among gilt litters. Gilts were randomized into test or control groups in a 1:1 ratio to compare the consumption of antibiotics used for piglets and piglet mortality during the four-week trial period. Control litters were treated with the oral antibiotic paromomycin, while test litters were treated with the complementary feed O-Nella-Protect. The farm trial included 18 gilt litters comprising 254 piglets. In the control group, 100% of the piglets received antibiotic treatment. In the test group, consumption of antibiotics used against diarrhea was reduced by 84% (p = 0.001) and consumption of antibiotics used for other illnesses was reduced by 45% (p = 0.045). In both test and control groups, six piglets died. Microbiological analysis identified both potential bacterial and viral pathogens. In conclusion, the farm trial indicates that even under the challenge of potentially serious bacterial and viral pathogens, a complimentary feed containing a tannin extract can support piglet health and reduce antibiotic consumption.

Single-Molecule Multivalent Interactions Revealed by Plasmon-Enhanced Fluorescence

Okholm, K. R., Nooteboom, S. W., Vinther, J. N., et al. — Sun, 01 Dec 2024 20:36:22 +0100

Multivalency as an interaction principle is widely utilized in nature. It enables specific and strong binding by multiple weak interactions through enhanced avidity and is a core process in immune recognition and cellular signaling, which is also a current concept in drug design. Here, we use the high signals from plasmon-enhanced fluorescence of nanoparticles to extract binding kinetics and dynamics of multivalent interactions on the single-molecule level and in real time. We study mono-, bi-, and trivalent binding interactions using a DNA Holliday Junction as a model construct with programmable valency and introduce a step-binding model for binding kinetics relevant for structured macromolecules by including an experimentally extractable binding restriction term ω to quantify the effects from conformation, steric effects, and rigidity. We used this approach to explore how length and flexibility of the DNA ligands affect binding restriction and binding strength, where the overall binding strength decreased with spacer length. For trivalent systems, increasing spacer length additionally activated binding in the trivalent state, giving insight into the design of multivalent drug or targeting moieties. By systematically changing the receptor density, we explored the binding super selectivity of the multivalent HJ at the single-molecule level. We find a polynomial behavior of the trivalent binding strength that clearly shows receptor-density-dependent selective binding. Interestingly, we could exploit the rapidly decaying near fields of the plasmon that induce a strong dependence of the signal on the position of the dye to observe binding dynamics during single multivalent binding events.

Shared genetic risk between anorexia nervosa and cardiovascular disease events

Qi, B., Graff, M., Bulik, C. M., North, K. E., Munn-Chernoff, M. A., Grove, J. — Thu, 01 Feb 2024 20:36:22 +0100

OBJECTIVE: Cardiovascular complications occur in up to 80% of patients with anorexia nervosa (AN), yet the underlying mechanisms warrant further investigation. We assessed the genetic correlation (rg ) between AN and cardiovascular disease (CVD) events to inform whether elevated cardiovascular risk among individuals with AN is due to shared genetic effects.

METHOD: We used genome-wide association study summary statistics for AN (N = 72,517), AN with binge eating (N = 12,630), AN without binge eating (N = 12,516), and six CVD events (N = 390,142 to 977,323). We calculated the rg s via linkage disequilibrium score regression and corrected for multiple testing using false discovery rate.

RESULTS: Significant rg s emerged between AN with heart failure (rg = -0.11, SE = 0.05, q = .04) and myocardial infarction (rg = -0.10, SE = 0.03, q = .01). AN with binge eating had a significant rg with myocardial infarction (rg = -0.15, SE = 0.06, q = .02). No significant rg emerged between AN without binge eating and any CVD event.

DISCUSSION: Some loci affect the liability to AN and CVD in opposite directions and the shared genetic effects may not be consistent across all CVD events. Our results provide further evidence suggesting that the elevated cardiovascular risk in AN may not be due to shared genetic underpinnings, but more likely a downstream consequence of the disease.

Structural and biochemical analysis of ligand binding in yeast Niemann–Pick type C1–related protein

Nel, L., Thaysen, K., Jamecna, D., et al. — Wed, 01 Jan 2025 20:36:22 +0100

In eukaryotes, integration of sterols into the vacuolar/lysosomal
membrane is critically dependent on the Niemann–Pick type C
(NPC) system. The system consists of an integral membrane
protein, called NCR1 in yeast, and NPC2, a luminal soluble protein
that transfers sterols to the N-terminal domain (NTD) of NCR1
before membrane integration. Both proteins have been implicated
in sterol homeostasis of yeast and humans. Here, we investigate
sterol and lipid binding of the NCR1/NPC2 transport
system and determine crystal structures of the sterol binding
NTD. The NTD binds both ergosterol and cholesterol, with nearly
identical conformations of the binding pocket. Apart from sterols,
the NTD can also bind fluorescent analogs of phosphatidylinositol,
phosphatidylcholine, and phosphatidylserine, as well as
sphingosine and ceramide. We confirm the multi-lipid scope of
the NCR1/NPC2 system using photo-crosslinkable and clickable
lipid analogs, namely, pac-cholesterol, pac-sphingosine, and pacceramide.
Finally, we reconstitute the transfer of pac-sphingosine
from NPC2 to the NTD in vitro. Collectively, our results support
that the yeast NPC system can work as versatile machinery for
vacuolar homeostasis of structurally diverse lipids, besides
ergosterol.

Analysis of exonic deletions in a large population study provides novel insights into NRXN1 pathology

Montalbano, S., Krebs, M. D., Rosengren, A., et al. — Sun, 01 Dec 2024 20:36:22 +0100

The NRXN1 locus is a hotspot for non-recurrent copy number variants and exon-disrupting NRXN1 deletions have been associated with increased risk of neurodevelopmental disorders in case-control studies. However, corresponding population-based estimates of prevalence and disease-associated risk are currently lacking. Also, most studies have not differentiated between deletions affecting exons of different NRXN1 splice variants nor considered intronic deletions. We used the iPSYCH2015 case-cohort sample to obtain unbiased estimates of the prevalence of NRXN1 deletions and their associated risk of autism, schizophrenia, depression, and ADHD. Most exon-disrupting deletions affected exons specific to the alpha isoform, and almost half of the non-exonic deletions represented a previously reported segregating founder deletion. Carriage of exon-disrupting NRXN1 deletions was associated with a threefold and twofold increased risk of autism and ADHD, respectively, whereas no significantly increased risk of depression or schizophrenia was observed. Our results highlight the importance of using population-based samples in genetic association studies.

Alkoxy Substituted Brominated closo-Dodecaborates with Functionalized Aliphatic Spacers

Sun, 01 Dec 2024 20:36:22 +0100

The utilization of dodecaborate boron clusters, [B₁₂X₁₂]²⁻ (X = Cl, Br, or I), as membrane carriers has been demonstrated recently, and their activity is known to be due to their superchaotropic nature. In this work, the mono-alkylation of [B₁₂Br₁₁OH]²⁻ to functionalize it with an aliphatic spacer was developed with a view to expanding the known chemical space of membrane carriers based on [B₁₂Br₁₂]²⁻. A new and improved facile route for the preparation of [B₁₂Br₁₁OH]²⁻, which is an important precursor to other [B₁₂Br₁₁OR]²⁻ species, is reported. Various alkoxylated [B₁₂Br₁₁O(CH₂)₅Z]²⁻ (Z = OH, N(CO)₂C₆H₄, CN and N₃) derivatives were prepared via a divergent synthesis based on [B₁₂Br₁₁O(CH₂)₅Br]²⁻. One of the newly synthesized compounds was utilized as a membrane carrier, and its impact on cell viability was examined.

Phosphine Oxide Indenoquinoline Derivatives

Rodriguez-Paniagua, A., Tesauro, C., Knudsen, B. R., Fuertes, M., Alonso, C. — Sun, 01 Dec 2024 20:36:22 +0100

The synthesis of phosphorous indenoquinolines and their biological evaluation as topoisomerase 1 (TOP1) inhibitors and antiproliferative agents were performed. First, the preparation of new hybrid 5H-indeno[2,1-c]quinolines with a phosphine oxide group was performed by a two-step Povarov-type [4+2]-cycloaddition reaction between the corresponding phosphorated aldimines with indene in the presence of BF₃·Et₂O. Subsequent oxidation of the methylene present in the structure resulted in the corresponding indeno[2,1-c]quinolin-7-one phosphine oxides 10. The synthesized derivatives were evaluated as TOP1 inhibitors showing higher inhibition values than CPT at prolonged incubation times (5 min). Inhibition of TOP1 was even observed after 30 min of incubation. The cytotoxic activities of these compounds were also studied against different cancer cell lines and a non-cancerous cell line. While some compounds showed cytotoxicity against some cancerous cells, none of the compounds showed any cytotoxicity against the non-cancerous cell line, MRC-5, in contrast to CPT, which exhibits high toxicity against this cell line. These results represent a very interesting advance since the heterocyclic phosphine oxide derivatives have important properties as TOP1 inhibitors and show an interesting cytotoxicity against different cell lines.

TrAnnoScope

Pektas, A., Panitz, F., Thomsen, B. — Sun, 01 Dec 2024 20:36:22 +0100

Background/Objectives: Transcriptome assembly and functional annotation are essential in understanding gene expression and biological function. Nevertheless, many existing pipelines lack the flexibility to integrate both short- and long-read sequencing data or fail to provide a complete, customizable workflow for transcriptome analysis, particularly for non-model organisms. Methods: We present TrAnnoScope, a transcriptome analysis pipeline designed to process Illumina short-read and PacBio long-read data. The pipeline provides a complete, customizable workflow to generate high-quality, full-length (FL) transcripts with broad functional annotation. Its modular design allows users to adapt specific analysis steps for other sequencing platforms or data types. The pipeline encompasses steps from quality control to functional annotation, employing tools and established databases such as SwissProt, Pfam, Gene Ontology (GO), the Kyoto Encyclopedia of Genes and Genomes (KEGG), and Eukaryotic Orthologous Groups (KOG). As a case study, TrAnnoScope was applied to RNA-Seq and Iso-Seq data from zebra finch brain, ovary, and testis tissue. Results: The zebra finch transcriptome generated by TrAnnoScope from the brain, ovary, and testis tissue demonstrated strong alignment with the reference genome (99.63%), and it was found that 93.95% of the matched protein sequences in the zebra finch proteome were captured as nearly complete. Functional annotation provided matches to known protein databases and assigned relevant functional terms to the majority of the transcripts. Conclusions: TrAnnoScope successfully integrates short and long sequencing technologies to generate transcriptomes with minimal user input. Its modularity and ease of use make it a valuable tool for researchers analyzing complex datasets, particularly for non-model organisms.

Dynamics of IGF Signaling During the Ovulatory Peak in Women Undergoing Ovarian Stimulation

Bøtkjær, J. A., Poulsen, L. l. C., Noer, P. R., et al. — Sun, 01 Dec 2024 20:36:23 +0100

CONTEXT: Insulin-like growth factor (IGF) signaling is known to affect human ovarian follicular function during growth and development. However, the role of the IGF system is unknown during the ovulatory peak, which is characterized by profound changes in granulosa cell (GCs) mitosis and function. OBJECTIVE: How is the IGF system expressed and regulated during the midcycle surge in women? METHODS: Follicular fluid (FF) and GCs were collected during the ovulatory peak from 2 specific time points. One sample was obtained before oocyte pickup (OPU): before ovulation trigger (OT) (T = 0 hours) or at 12, 17, or 32 hours after OT, and 1 sample was obtained at OPU 36 hours after OT. Fifty women undergoing ovarian stimulation at a university hospital were included. Gene expression profiles were assessed by microarray analysis of GCs. IGF-related proteins in the FF were assessed by immunoassay or by determination of activity with a proteinase assay. RESULTS: Gene expression of proteins promoting IGF activity (ie, IGF2, PAPP-A, and IRS1) together with proliferation markers were downregulated on a transcriptional level in GCs after OT, whereas proteins inhibiting the IGF signal (ie, IGFBPs, IGF2, and STC1) were upregulated. STC1 gene expression and protein levels were greatly upregulated after OT with a parallel steep downregulation of PAPP-A proteolytic activity. CONCLUSION: These data suggest that downregulation of IGF signaling mediated by increased STC1 expression is instrumental for the sudden cessation in GC proliferation and onset of differentiation during the ovulatory peak.

The Cryo-EM structure of human CD163 bound to haptoglobin-hemoglobin reveals molecular mechanisms of hemoglobin scavenging

Etzerodt, A., Mikkelsen, J. H., Torvund-Jensen, M., et al. — Mon, 30 Dec 2024 20:36:23 +0100

CD163, a macrophage-specific receptor, plays a critical role in scavenging hemoglobin released during hemolysis, protecting against oxidative effects of heme iron. In the bloodstream, hemoglobin is bound by haptoglobin, leading to its immediate endocytosis by CD163. While haptoglobin's structure and function are well understood, CD163's structure and its interaction with the haptoglobin-hemoglobin complex have remained elusive. Here, we present the cryo-electron microscopy structure of the entire extracellular domain of human CD163 in complex with haptoglobin-hemoglobin. The structure reveals that CD163 assembles into trimers (and to some extent dimers), binding haptoglobin-hemoglobin in their center. Key acidic residues in CD163 interact with lysine residues from both haptoglobin and hemoglobin. Calcium-binding sites located near the haptoglobin-hemoglobin interface in CD163 provide explanation for the calcium dependence of the interaction. Furthermore, we show that the interaction facilitating CD163 oligomerization mimics ligand binding and is also calcium dependent. This structural insight into CD163 advances our understanding of its role in hemoglobin scavenging as well as its broader relevance to structurally related scavenger receptors.

Optimized Construction of a Yeast SICLOPPS Library for Unbiased In Vivo Selection of Cyclic Peptides

Birkmose, N., Frydendahl, E. U., Knudsen, C. R. — Sun, 01 Dec 2024 20:36:23 +0100

DNA-encoded libraries hold great potential for discovering small, cyclized peptides with drug potential. Split-intein circular ligation of peptides and proteins (SICLOPPS) is a well-established method for in vivo selection of cyclic peptides targeting specific intracellular components. However, the method has mainly been used in prokaryotic cells. In contrast, selection studies performed directly in eukaryotic cells allow for the identification of cyclic peptides promoting a functional outcome, without the need to define a specific cellular target. Here, we report the construction of a Saccharomyces cerevisiae-specific SICLOPPS library of 80 million members, via careful optimization of several steps to increase the size of the library. Individual library members were shown to be correctly expressed and processed in yeast. High-throughput sequencing was conducted on the randomized primer used for library construction and the pure yeast SICLOPPS library isolated from Escherichia coli. A distinct guanine insertion bias was observed in the peptide-encoding, randomized sequence, which was primarily attributed to the degenerate primer used to introduce the randomized sequence. Moreover, high-throughput sequencing was performed on the library before and after the induction of cyclic peptide expression in yeast. Importantly, expression of the SICLOPPS library in S. cerevisiae caused only a marginal further sequence bias. Our work paves the way for selection studies using a large and diverse library to identify cyclic peptides of therapeutic interest that promote a specific phenotypic outcome in eukaryotic organisms, with yeast representing a beneficial model system due to its high transformation efficiency.

Resilience and Charge-Dependent Fibrillation of functional amyloid

Golan, N., Parizat, A., Tabachnikov, O., et al. — Sat, 01 Feb 2025 20:36:23 +0100

FapC and FapB are biofilm-associated amyloids involved in the virulence of Pseudomonas and other bacteria. We herein demonstrate their exceptional thermal and chemical resilience, suggesting that their biofilm structures might withstand standard sterilization, thereby contributing to the persistence of Pseudomonas aeruginosa infections. Our findings also underscore the impact of environmental factors on functional amyloid in Pseudomonas (Fap) proteins, suggesting that orthologs in different Pseudomonas strains adapt to specific environments and roles. Challenging previous assumptions about a simple nucleation role for FapB in promoting FapC aggregation, the study shows a significant influence of FapC on FapB aggregation. The interaction between these FapB and FapC is intricate: FapB stabilizes FapC fibrils, while FapC slows down FapB fibrillation but can still serve as a cross-seeding template. This complex interplay is the key to understanding their roles in bacterial biofilms. Furthermore, the study highlights distinct differences between Fap and Escherichia coli's CsgA (curli) amyloid, where CsgB assumes a simple unidirectional role in nucleating CsgA fibrillation, emphasizing the importance of a comprehensive understanding of various amyloid systems. This knowledge is vital for developing effective intervention strategies against bacterial infections and leveraging the unique properties of these amyloids in technological applications such as novel bionanomaterials or protective coatings.

Modeling of mRNA deadenylation rates reveal a complex relationship between mRNA deadenylation and decay

Czarnocka-Cieciura, A., Poznański, J., Turtola, M., et al. — Sun, 01 Dec 2024 20:36:23 +0100

Complete cytoplasmic polyadenosine tail (polyA-tail) deadenylation is thought to be essential for initiating mRNA decapping and subsequent degradation. To investigate this prevalent model, we conducted direct RNA sequencing of S. cerevisiae mRNAs derived from chase experiments under steady-state and stress condition. Subsequently, we developed a numerical model based on a modified gamma distribution function, which estimated the transcriptomic deadenylation rate at 10 A/min. A simplified independent method, based on the delineation of quantile polyA-tail values, showed a correlation between the decay and deadenylation rates of individual mRNAs, which appeared consistent within functional transcript groups and associated with codon optimality. Notably, these rates varied during the stress response. Detailed analysis of ribosomal protein-coding mRNAs (RPG mRNAs), constituting 40% of the transcriptome, singled out this transcript group. While deadenylation and decay of RPG mRNAs accelerated under heat stress, their degradation could proceed even when deadenylation was blocked, depending entirely on ongoing nuclear export. Our findings support the general primary function of deadenylation in dictating the onset of decapping, while also demonstrating complex relations between these processes.

Rescuable sleep and synaptogenesis phenotypes in a Drosophila model of O-GlcNAc transferase intellectual disability

Fri, 01 Nov 2024 20:36:23 +0100

O-GlcNAcylation is an essential intracellular protein modification mediated by O-GlcNAc transferase (OGT) and O-GlcNAcase (OGA). Recently, missense mutations in OGT have been linked to intellectual disability, indicating that this modification is important for the development and functioning of the nervous system. However, the processes that are most sensitive to perturbations in O-GlcNAcylation remain to be identified. Here, we uncover quantifiable phenotypes in the fruit fly Drosophila melanogaster carrying a patient-derived OGT mutation in the catalytic domain. Hypo-O-GlcNAcylation leads to defects in synaptogenesis and reduced sleep stability. Both these phenotypes can be partially rescued by genetically or chemically targeting OGA, suggesting that a balance of OGT/OGA activity is required for normal neuronal development and function.

Exercise, hormesis and ageing

Radak, Z., Rattan, S. I. S. — Sat, 01 Feb 2025 20:36:23 +0100

Non-Invasive Malaria Detection in Sub-Saharan Africa Using a DNA-Based Sensor System

Juul-Kristensen, T., Thiesen, C., Haurum, L. W., et al. — Sun, 01 Dec 2024 20:36:23 +0100

Malaria poses a serious global health problem, with half the world population being at risk. Regular screening is crucial for breaking the transmission cycle and combatting the disease spreading. However, current diagnostic tools relying on blood samples face challenges in many malaria-epidemic areas. In the present study, we demonstrate the detection of the malaria-causing Plasmodium parasite in non-invasive saliva samples (N = 61) from infected individuals by combining a DNA-based Rolling-circle-Enhanced-Enzyme-Activity-Detection (REEAD) sensor system with a chemiluminescence readout that could be detected with an in-house-developed affordable and battery-powered portable reader. We successfully transferred the technology to sub-Saharan Africa, where the malaria burden is high, and demonstrated a proof of concept in a small study (N = 40) showing significant differences (p < 0.00001) between malaria-positive individuals (N = 33) and presumed asymptomatic negative individuals (N = 7) all collected in Gabon. This is the first successful application of the REEAD sensor system for the detection of malaria in saliva in a high-epidemic area and holds promise for the potential future use of REEAD for malaria diagnosis or surveillance based on non-invasive specimens in sub-Saharan Africa.

DNA-directed termination of mammalian RNA polymerase II

Davidson, L., Rouvière, J. O., Sousa-Luís, R., et al. — Fri, 01 Nov 2024 20:36:23 +0100

The best-studied mechanism of eukaryotic RNA polymerase II (RNAPII) transcriptional termination involves polyadenylation site-directed cleavage of the nascent RNA. The RNAPII-associated cleavage product is then degraded by XRN2, dislodging RNAPII from the DNA template. In contrast, prokaryotic RNAP and eukaryotic RNAPIII often terminate directly at T-tracts in the coding DNA strand. Here, we demonstrate a similar and omnipresent capability for mammalian RNAPII. Importantly, this termination mechanism does not require upstream RNA cleavage. Accordingly, T-tract-dependent termination can take place when XRN2 cannot be engaged. We show that T-tracts can terminate snRNA transcription independently of RNA cleavage by the Integrator complex. Importantly, we found genome-wide termination at T-tracts in promoter-proximal regions but not within protein-coding gene bodies. XRN2-dependent termination dominates downstream from protein-coding genes, but the T-tract process is sometimes used. Overall, we demonstrate global DNA-directed attrition of RNAPII transcription, suggesting that RNAPs retain the potential to terminate over T-rich sequences throughout evolution.

RNA 3′end tailing safeguards cells against products of pervasive transcription termination

Wu, G., Rouvière, J. O., Schmid, M., Heick Jensen, T. — Sun, 01 Dec 2024 20:36:23 +0100

Premature transcription termination yields a wealth of unadenylated (pA⁻) RNA. Although this can be targeted for degradation by the Nuclear EXosome Targeting (NEXT) complex, possible backup pathways remain poorly understood. Here, we find increased levels of 3′ end uridylated and adenylated RNAs upon NEXT inactivation. U-tailed RNAs are mostly short and modified by the cytoplasmic tailing enzymes, TUT4/7, following their PHAX-dependent nuclear export and prior to their degradation by the cytoplasmic exosome or the exoribonuclease DIS3L2. Longer RNAs are instead adenylated redundantly by enzymes TENT2, PAPOLA and PAPOLG. These transcripts are either degraded via the nuclear Poly(A) tail eXosome Targeting (PAXT) connection or exported and removed by the cytoplasmic exosome in a translation-dependent manner. Failure to do so decreases global translation and induces cell death. We conclude that post-transcriptional 3′ end modification and removal of excess pA⁻ RNA is achieved by tailing enzymes and export factors shared with productive RNA pathways.

Increased diversity of beneficial rhizobia enhances faba bean growth

Mendoza-Suárez, M., Akyol, T. Y., Nadzieja, M., Andersen, S. U. — Sun, 01 Dec 2024 20:36:23 +0100

Legume-rhizobium symbiosis provides a sustainable nitrogen source for agriculture. Nitrogen fixation efficiency depends on both legume and rhizobium genotypes, but the implications of their interactions for plant performance in environments with many competing rhizobium strains remain unclear. Here, we let 399 Rhizobium leguminosarum complex sv. viciae strains compete for nodulation of 212 faba bean genotypes. We find that the strains can be categorised by their nodule occupancy profiles into groups that show distinct competitive interactions and plant growth-promoting effects. Further, we show that the diversity of strains occupying root nodules affects plant growth and is under plant genetic control. These insights provide a basis for re-designing rhizobium inoculation and plant breeding strategies to enhance symbiotic nitrogen fixation in agriculture.

A cautionary tale for Alzheimer’s disease GWAS by proxy

Pedersen, E. M., Wimberley, T., Vilhjálmsson, B. J. — Mon, 02 Dec 2024 20:36:23 +0100

Using reported parental disease history to decipher the genetics of Alzheimer’s disease may be promising, but this approach is also susceptible to complex selection and information bias that can mislead researchers if not accounted for.

Enhancing Membrane Permeability of Fluorescein-Type Chromophore Through Covalent Attachment of Chlorinated Dodecaborate

Fri, 01 Nov 2024 20:36:23 +0100

Anionic boron clusters, such as [B₁₂X₁₂]²⁻ (X = Cl, Br, I), have attracted attention in pharmaceuticals due to their unique superchaotropic properties. In particular, [B₁₂Br₁₂]²⁻ (1) has demonstrated strong interactions with biomolecules, facilitating cargo translocation across plasma membranes. In this study, we investigated the effect of covalently attaching chlorinated dodecaborate moiety [B₁₂Cl₁₁O-]²⁻ to 6-carboxyfluorescein (6-FAM) (3) via a PEG3 linker to form conjugate (4). We compared the membrane permeability of this covalent conjugate with that of non-covalent interactions between 6-FAM (3) and [B₁₂Cl₁₂]²⁻ (2). Live-cell fluorescence imaging revealed that the covalent conjugate exhibited enhanced membrane permeability and water solubility while maintaining low cytotoxicity. These results highlight the potential of covalent conjugation with boron clusters for improving the cellular uptake of hydrophilic cargos.

Dimethyl labeling of N-terminal amines allows unambiguous identification of protein crosslinks

Sat, 01 Feb 2025 20:36:23 +0100

Protein crosslinks induced through either deliberate enzymatic oxidation or reactive oxidants (oxidative eustress/distress), are associated with multiple human pathologies including atherosclerosis, Alzheimer's and Parkinson's diseases. In many cases, the nature of the crosslinks, their position(s) either within (intramolecular) or between (intermolecular) polypeptide chains, and concentrations are unclear. Although limited data are available from specific antibodies, detailed characterization of protein crosslinks is often performed by mass spectrometric analysis of peptides from proteolytic digestion. Such analyses are challenging due to the low concentration of these species, and the complexity of their fragment ion spectra when compared to non-crosslinked species. We hypothesized that highly efficient and specific chemical amine labeling of the two N-termini in crosslinked peptides (compared to the single N-terminus of linear peptides), using “light” and “heavy” isotope-labelled reagents would facilitate identification, validation and quantification of crosslinks. This method was compared to a previous enzyme-catalyzed ¹⁸O C-terminal carboxylate labeling approach. N-terminal amine dimethyl labeling is shown to have major advantages over the ¹⁸O-approach including high labeling yields (92–100 %) and well-defined mass spectrometric isotope distribution patterns. This approach has allowed identification of novel dityrosine crosslinks between pair of tyrosine (Tyr, Y) residues in photo-oxidized β-casein (Y195-Y195, Y195-Y208, Y208-Y208), and α-synuclein exposed to nitrosative stress (Y39-Y39, Y39-Y125, Y39-Y133, Y133-Y136). This approach is also applicable to disulfide bond mapping, with 15 of 17 disulfides in serum albumin readily detected. These data indicate that dimethyl labeling is a highly versatile and efficient approach for the site-specific identification of oxidation- and nitration-induced crosslinks in proteins.

Nanobodies' duo facilitates ultrasensitive serum HER-2/neu immunoassays via enhanced avidity interactions

Boonkaew, S., Teodori, L., Vendelbo, M. H., Kjems, J., Ferapontova, E. E. — Wed, 15 Jan 2025 20:36:23 +0100

BACKGROUND: Existing liquid biopsy assays for protein biomarkers of cancer are mostly based on antibodies (Ab) contributing unfavorably to their high cost. Easy to express and modify in vitro, nanobodies may be a cost-effective alternative to Ab.

RESULTS: We show that serum HER-2/neu, a biomarker and target of aggressive HER-2/neu(+) cancers, can be accurately detected in a 1.2 h electrochemical cellulase-linked sandwich nanobody/aptamer assay on magnetic beads. Using a single nanobody receptor, 2Rs15d or 2Rb17c, reduces immunoassay's sensitivity by 35%-26 %. A combination of two nanobodies as a duo-receptor recovers the sensitivity of the enzyme-linked nanobody/aptamer-sorbent assay (ELNASA) to 11.9 ± 2.8 μC fM-1, due to the avidity effects making the nanobodies-duo binding properties comparable to those of Ab. Down to 0.1 fM HER-2/neu was detected by ELNASA in serum samples, with no interference from other blood-circulating proteins. In a 30 healthy-volunteers trial, ELNASA more accurately than optical ELISA assayed serum HER-2/neu.

SIGNIFICANCE: ELNASA performance rivals that of ELISA, yet estimated to be at least 200 times cheaper, due to the lower cost of nanobodies production, and may be better suited for routine clinical analysis of HER-2/neu, particularly, in low- and middle-income settings with limited resources. The ELNASA approach is generic and may be adapted for specific and ultrasensitive analysis of other blood-circulating proteins.

Distinct fingerprints of tRNA-derived small non-coding RNA in animal models of neurodegeneration

Baindoor, S., Gibriel, H. A.Y., Venø, M. T., et al. — Fri, 01 Nov 2024 20:36:23 +0100

Transfer RNA-derived small RNAs (tsRNAs) - categorized as tRNA-derived fragments (tRFs), tRNA-derived stress-induced RNAs (tiRNAs) and internal tRF (itRF) - are small non-coding RNAs that participate in various cellular processes such as translation inhibition and responses to cellular stress. We here identified tsRNA profiles within susceptible tissues in animal models of amyotrophic lateral sclerosis (ALS), frontotemporal dementia (FTD) and Parkinson's disease (PD) to pinpoint disease-specific tsRNAs and those shared across neurodegenerative diseases. We performed small RNA sequencing in the SOD1G93A and TDP43A315T mouse models of ALS (spinal cord), the TauP301S model of FTD (hippocampus), and the parkin/POLG model of PD (substantia nigra). Bioinformatic analysis showed higher expression of 5' tiRNAs selectively in the two ALS models, lower expression of 3' tRFs in both the ALS and FTD mouse models, and lower expression of itRF Arg in the PD model. Experimental validation confirmed the expression of tsRNAs. Gene Ontology analysis of targets associated with validated 3' tRFs indicated functions in the regulation of synaptic and neuronal pathways. Our profiling of tsRNAs indicates disease-specific fingerprints in animal models of neurodegeneration, which require validation in human disease.

Young rat microbiota extracts strongly inhibit fibrillation of α-synuclein and protect neuroblastoma cells and zebrafish against α-synuclein toxicity

Shiraz, M. G., Nielsen, J., Widmann, J., et al. — Wed, 01 Jan 2025 20:36:23 +0100

The clinical manifestations of Parkinson's disease (PD) are driven by aggregation of α-Synuclein (α-Syn) in the brain. However, there is increasing evidence that PD may be initiated in the gut and thence spread to the brain, eg, via the vagus nerve. Many studies link PD to changes in the gut microbiome, and bacterial amyloid has been shown to stimulate α-Syn aggregation. Yet, we are not aware of any studies reporting on a direct connection between microbiome components and α-Syn aggregation. Here, we report that soluble extract from the gut microbiome of the rats, particularly young rats transgenic for PD, shows a remarkably strong ability to inhibit in vitro α-Syn aggregation and keep it natively unfolded and monomeric. The active component(s) are heat-labile molecule(s) of around 30- to 100-kDa size, which are neither nucleic acid nor lipid. Proteomic analysis identified several proteins whose concentrations in different rat samples correlated with the samples’ anti-inhibitory activity, while a subsequent pull-down assay linked the protein chaperone DnaK with the inhibitory activity of young rat's microbiome, confirmed in subsequent in vitro assays. Remarkably, the microbiome extracts also protected neuroblastoma SH-SY5Y cells and zebrafish embryos against α-Syn toxicity. Our study sheds new light on the gut microbiome as a potential source of protection against PD and opens up for new microbiome-based therapeutic strategies.

m⁶A modification of mutant huntingtin RNA promotes the biogenesis of pathogenic huntingtin transcripts

Pupak, A., Rodríguez-Navarro, I., Sathasivam, K., et al. — Fri, 08 Nov 2024 20:36:23 +0100

In Huntington’s disease (HD), aberrant processing of huntingtin (HTT) mRNA produces HTT1a transcripts that encode the pathogenic HTT exon 1 protein. The mechanisms behind HTT1a production are not fully understood. Considering the role of m⁶A in RNA processing and splicing, we investigated its involvement in HTT1a generation. Here, we show that m⁶A methylation is increased before the cryptic poly(A) sites (IpA1 and IpA2) within the huntingtin RNA in the striatum of Hdh+/Q111 mice and human HD samples. We further assessed m⁶A’s role in mutant Htt mRNA processing by pharmacological inhibition and knockdown of METTL3, as well as targeted demethylation of Htt intron 1 using a dCas13-ALKBH5 system in HD mouse cells. Our data reveal that Htt1a transcript levels are regulated by both METTL3 and the methylation status of Htt intron 1. They also show that m⁶A methylation in intron 1 depends on expanded CAG repeats. Our findings highlight a potential role for m⁶A in aberrant splicing of Htt mRNA.

Hydrogel bead-based isothermal detection (BEAD-ID) for assessing the activity of DNA-modifying enzymes

Borg, K. N., Shetty, A., Cheng, G., et al. — Fri, 20 Dec 2024 20:36:23 +0100

DNA-modifying enzymes are crucial in biological processes and have significant clinical implications. Traditional quantification methods often overlook enzymatic activity, the true determinants of enzymes’ functions. We present hydrogel Bead-based Isothermal Detection (BEAD-ID), utilizing uniform hydrogel bead-based microreactors to evaluate DNA-modifying enzyme activity on-bead. We fabricated homogeneous oligo-conjugated polyacrylamide (oligo-PAA) beads via droplet microfluidics, optimized for capturing and amplifying enzyme-modified nanosensors. By incorporating DNA oligos within the hydrogel network, BEAD-ID retains isothermally amplified products, facilitating in situ detection of enzyme activities on-bead. We validate BEAD-ID by quantifying human topoisomerase I (TOP1) and restriction endonuclease EcoRI, showing a direct correlation between enzyme concentration and fluorescence intensity, demonstrating the platform's sensitivity (6.25 nM TOP1, 6.25 U/μL EcoRI) and reliability in food matrix (25 U/μL EcoRI). Additionally, a customized flow cytometry-mimicking setup allows high-throughput detection at 352 Hz with objective assessment. BEAD-ID, offering flexibility and scalability, is a promising tool for studying DNA-modifying enzymes.

Hydrogel bead-based isothermal detection (BEAD-ID) for assessing the activity of DNA-modifying enzymes

Borg, K. N., Shetty, A., Cheng, G. ., et al. — Fri, 20 Dec 2024 20:36:23 +0100

Compartment-specific small non-coding RNA changes and nucleolar defects in human mesial temporal lobe epilepsy

Vangoor, V. R., Giuliani, G., de Wit, M., et al. — Sun, 01 Dec 2024 20:36:23 +0100

Mesial temporal lobe epilepsy (mTLE) is a debilitating disease characterized by recurrent seizures originating from temporal lobe structures such as the hippocampus. The pathogenic mechanisms underlying mTLE are incompletely understood but include changes in the expression of non-coding RNAs in affected brain regions. Previous work indicates that some of these changes may be selective to specific sub-cellular compartments, but the full extent of these changes and how these sub-cellular compartments themselves are affected remains largely unknown. Here, we performed small RNA sequencing (RNA-seq) of sub-cellular fractions of hippocampal tissue from mTLE patients and controls to determine nuclear and cytoplasmic expression levels of microRNAs (miRNAs). This showed differential expression of miRNAs and isomiRs, several of which displayed enriched nuclear expression in mTLE. Subsequent analysis of miR-92b, the most strongly deregulated miRNA in the nucleus, showed accumulation of this miRNA in the nucleolus in mTLE and association with snoRNAs. This prompted us to further study the nucleolus in human mTLE which uncovered several defects, such as altered nucleolar size or shape, mis-localization of nucleolar proteins, and deregulation of snoRNAs, indicative of nucleolar stress. In a rat model of epilepsy, nucleolar phenotypes were detected in the latency period before the onset of spontaneous seizures, suggesting that nucleolar changes may contribute to the development of seizures and mTLE. Overall, these data for the first time implicate nucleolar defects in the pathogenesis of mTLE and provide a valuable framework for further defining the functional consequences of altered sub-cellular RNA profiles in this disease.

A conserved juxtamembrane motif in plant NFR5 receptors is essential for root nodule symbiosis

Hansen, S. B., Luu, T. B., Gysel, K., et al. — Fri, 01 Nov 2024 20:36:23 +0100

Establishment of root nodule symbiosis is initiated by the perception of bacterial Nod factor ligands by the plant LysM receptor kinases NFR1 and NFR5. Receptor signaling initiating the symbiotic pathway depends on the kinase activity of NFR1, while the signaling mechanism of the catalytically inactive NFR5 pseudokinase is unknown. Here, we present the crystal structure of the signaling-competent Lotus japonicus NFR5 intracellular domain, comprising the juxtamembrane region and pseudokinase domain. The juxtamembrane region is structurally well defined and forms two α-helices, αA and αA', which contain an exposed hydrophobic motif. We demonstrate that this "juxtamembrane motif" promotes NFR5-NFR5 and NFR1-NFR5 interactions and is essential for symbiotic signaling. Conservation analysis reveals that the juxtamembrane motif is present throughout NFR5-type receptors and is required for symbiosis signaling from barley RLK10, suggesting a conserved and broader function for this motif in plant-microbe symbioses.

Magnetoresponsive liposomes applications in nanomedicine

Shahsavari, S., Rad, M. B., Hajiaghajani, A., et al. — Sun, 01 Dec 2024 20:36:23 +0100

Safe and effective cancer therapy requires a suitable nanocarrier that can target particular sites, such as cancer cells, in a selective manner. With the tremendous growth in nanotechnology, liposomes, among various competing nanocarriers, have shown promising advances in cancer therapy. Magnetic nanoparticles and metal ions are wide-reaching candidates for conferring magnetic properties and for incorporation into liposomes. Combining liposomes with magnetic structures enables construction of magnetoresponsive liposomes, allowing stimuli-responsiveness to an alternating magnetic field, magnetic targeting, and tracking by magnetic resonance imaging, which could all occur in parallel. This review presents a comprehensive analysis of the practical advances and novel aspects of design, synthesis and engineering magnetoresponsive liposomes, emphasizing their diverse properties for various applications. Our work explores the innovative uses of these structures, extending beyond drug delivery to include smart contrast agents, cell labeling, biosensing, separation, and filtering. By comparing new findings with earlier studies, we showcase significant improvements in efficiency and uncover new potentials, setting a new benchmark for future research in the field of magnetoresponsive liposomes.

Extensive Population Structure Highlights an Apparent Paradox of Stasis in the Impala (Aepyceros melampus)

Garcia-Erill, G., Wang, X., Rasmussen, M., et al. — Fri, 01 Nov 2024 20:36:23 +0100

Impalas are unusual among bovids because they have remained morphologically similar over millions of years—a phenomenon referred to as evolutionary stasis. Here, we sequenced 119 whole genomes from the two extant subspecies of impala, the common (Aepyceros melampus melampus) and black-faced (A. m. petersi) impala. We investigated the evolutionary forces working within the species to explore how they might be associated with its evolutionary stasis as a taxon. Despite being one of the most abundant bovid species, we found low genetic diversity overall, and a phylogeographic signal of spatial expansion from southern to eastern Africa. Contrary to expectations under a scenario of evolutionary stasis, we found pronounced genetic structure between and within the two subspecies with indications of ancient, but not recent, gene flow. Black-faced impala and eastern African common impala populations had more runs of homozygosity than common impala in southern Africa, and, using a proxy for genetic load, we found that natural selection is working less efficiently in these populations compared to the southern African populations. Together with the fossil record, our results are consistent with a fixed-optimum model of evolutionary stasis, in which impalas in the southern African core of the range are able to stay near their evolutionary fitness optimum as a generalist ecotone species, whereas eastern African impalas may struggle to do so due to the effects of genetic drift and reduced adaptation to the local habitat, leading to recurrent local extinction in eastern Africa and re-colonisation from the South.

DiDBiT-TMT

Gamaleldin, M., Yu, N. K., Diedrich, J. K., et al. — Fri, 01 Nov 2024 20:36:23 +0100

Direct detection of biotinylated proteins (DiDBiT) is a proteomic method that can enrich and detect newly synthesized proteins (NSPs) labeled with bio-orthogonal amino acids with 20-fold improved detectability compared to conventional methods. However, DiDBiT has currently been used to compare only two conditions per experiment. Here, we present DiDBiT-TMT, a method that can be used to quantify NSPs across many conditions and replicates in the same experiment by combining isobaric tandem mass tagging (TMT) with DiDBiT. We applied DiDBiT-TMT to brain slices to determine changes in the de novo proteome that occur after inducing chemical long-term potentiation (cLTP) or treatment with the neuromodulator norepinephrine. We successfully demonstrated DiDBiT-TMT’s capacity to quantitatively compare up to 9 samples in parallel. We showed that there is a minimal overlap among NSPs that are differentially expressed in cLTP-treated organotypic brain slices, norepinephrine-treated organotypic brain slices, and organotypic slices undergoing combinatorial treatment with norepinephrine and cLTP. Our results point to the possible divergence of the molecular mechanisms underlying these treatments and showcase the applicability of DiDBiT-TMT for studying neurobiology.

Natural variation in root exudate composition in the genetically structured Arabidopsis thaliana in the Iberian Peninsula

Sat, 01 Feb 2025 20:36:23 +0100

Genetic liability estimated from large-scale family data improves genetic prediction, risk score profiling, and gene mapping for major depression

Dybdahl Krebs, M., Georgii Hellberg, K. L., Lundberg, M., et al. — Thu, 07 Nov 2024 20:36:23 +0100

Large biobank samples provide an opportunity to integrate broad phenotyping, familial records, and molecular genetics data to study complex traits and diseases. We introduce Pearson-Aitken Family Genetic Risk Scores (PA-FGRS), a method for estimating disease liability from patterns of diagnoses in extended, age-censored genealogical records. We then apply the method to study a paradigmatic complex disorder, major depressive disorder (MDD), using the iPSYCH2015 case-cohort study of 30,949 MDD cases, 39,655 random population controls, and more than 2 million relatives. We show that combining PA-FGRS liabilities estimated from family records with molecular genotypes of probands improves three lines of inquiry. Incorporating PA-FGRS liabilities improves classification of MDD over and above polygenic scores, identifies robust genetic contributions to clinical heterogeneity in MDD associated with comorbidity, recurrence, and severity and can improve the power of genome-wide association studies. Our method is flexible and easy to use, and our study approaches are generalizable to other datasets and other complex traits and diseases.

The effects of Zanthoxylum bungeanum extract on lipid metabolism induced by sterols

Wu, T., Zhong, L., Hong, Z., et al. — Sun, 01 Mar 2015 20:36:23 +0100

Variant pharmacological activities of Zanthoxylum bungeanum were determined before. The aim of this study was to assess whether Z. bungeanum could regulate lipid metabolism. The cholesterol overloading HepG2 cells induced by sterols were used as in vitro model to study lipid-lowering activities of the n-butanol (BuOH) fraction isolated from Z. bungeanum (ZBBu). Male apolipoprotein E knockout (apoE-KO) mice with high fat diet were used as in vivo model. We firstly demonstrated ZBBu had effects on reversed lipid accumulation, decreased apoB and enhanced apoA1 secretion. It increased the amount of low density lipoprotein receptor (LDLR) protein, also significantly inhibited the expression of SREBP-1 and SREBP-2's target molecule (hydroxy methylglutaryl coenzyme A reductase, HMGCR), which might be active in stimulation of RCT. And the expression of genes involved in RCT, such as CYP27A1, LXR-α, ABCG1, was promoted by ZBBu. Furthermore, ZBBu could reduce serum TC, TG levels in apoE-KO mice. Our study indicated that ZBBu could regulate the lipid metabolism through increasing the amount of low density lipoprotein receptor (LDLR) and inducing the expression of genes involved in RCT.

Novel metabolic role for BDNF in pancreatic β-cell insulin secretion

Fulgenzi, G., Hong, Z., Tomassoni-Ardori, F., et al. — Tue, 01 Dec 2020 20:36:23 +0100

BDNF signaling in hypothalamic circuitries regulates mammalian food intake. However, whether BDNF exerts metabolic effects on peripheral organs is currently unknown. Here, we show that the BDNF receptor TrkB.T1 is expressed by pancreatic β-cells where it regulates insulin release. Mice lacking TrkB.T1 show impaired glucose tolerance and insulin secretion. β-cell BDNF-TrkB.T1 signaling triggers calcium release from intracellular stores, increasing glucose-induced insulin secretion. Additionally, BDNF is secreted by skeletal muscle and muscle-specific BDNF knockout phenocopies the β-cell TrkB.T1 deletion metabolic impairments. The finding that BDNF is also secreted by differentiated human muscle cells and induces insulin secretion in human islets via TrkB.T1 identifies a new regulatory function of BDNF on metabolism that is independent of CNS activity. Our data suggest that muscle-derived BDNF may be a key factor mediating increased glucose metabolism in response to exercise, with implications for the treatment of diabetes and related metabolic diseases.

Generation of Functional Mouse Hippocampal Neurons

Tomassoni-Ardori, F., Hong, Z., Fulgenzi, G., Tessarollo, L. — Wed, 05 Aug 2020 20:36:23 +0200

Primary culture of mouse hippocampal neurons is a very useful in vitro model for studying neuronal development, axonal and dendritic morphology, synaptic functions, and many other neuronal features. Here we describe a step-by-step process of generating primary neurons from mouse embryonic hippocampi (E17.5/E18.5). Hippocampal neurons generated with this protocol can be plated in different tissue culture dishes according to different experimental aims and can produce a reliable source of pure and differentiated neurons in less than one week. This protocol covers all the steps necessary for the preparation, culture and characterization of the neuronal culture, including the illustration of dissection instruments, surgical procedure for embryos' isolation, culturing conditions and assessment of culture's purity and differentiation. Evaluation of neuronal activity was performed by analysis of calcium imaging dynamics at six days in culture.

TrkA-cholinergic signaling modulates fear encoding and extinction learning in PTSD-like behavior

Yanpallewar, S., Tomassoni-Ardori, F., Palko, M. E., et al. — Thu, 01 Dec 2022 20:36:23 +0100

Recent studies have suggested that the use of cognitive enhancers as adjuncts to exposure-based therapy in individuals suffering from post-traumatic stress disorder (PTSD) may be beneficial. Brain cholinergic signaling through basal forebrain projections to the hippocampus is an established pathway mediating fear response and cognitive flexibility. Here we employed a genetic strategy to enhance cholinergic activity through increased signaling of the NGF receptor TrkA. This strategy leads to increased levels of the marker of cholinergic activation, acetylcholine synthesizing enzyme choline acetyltransferase, in forebrain cholinergic regions and their projection areas such as the hippocampus. Mice with increased cholinergic activity do not display any neurobehavioral abnormalities except a selective attenuation of fear response and lower fear expression in extinction trials. Reduction in fear response is rescued by the GABA antagonist picrotoxin in mutant mice, and, in wild-type mice, is mimicked by the GABA agonist midazolam suggesting that GABA can modulate cholinergic functions on fear circuitries. Importantly, mutant mice also show a reduction in fear processing under stress conditions in a single prolonged stress (SPS) model of PTSD-like behavior, and augmentation of cholinergic signaling by the drug donepezil in wild-type mice promotes extinction learning in a similar SPS model of PTSD-like behavior. Donepezil is already in clinical use for the treatment of dementia suggesting a new translational application of this drug for improving exposure-based psychotherapy in PTSD patients.

Fat supplement for dairy cows during early lactation – potentials, challenges, and risks – a meta-analysis

Lashkari, S., Weisbjerg, M. R., Foldager, L., Børsting, C. F. — Fri, 01 Mar 2024 20:36:23 +0100

The most critical challenge in early lactation is to meet the increasing energy demand for milkproduction, which results in a negative energy balance (EB). The objective of this study was toevaluate the productive performance and plasma metabolite responses of early lactating dairy cowsfed diets supplemented with increasing levels of different fat sources. Diets were categorized as dietssupplemented with saturated fatty acid rich fat sources (SFAR), with Ca-soaps of fatty acids (CaFA),and with polyunsaturated fatty acid rich fat sources (PUFAR). There was an interaction between crudefat level and source for dry matter intake (DMI); each percentage-unit increase reduced DMI by 0.38and 0.77 kg/d in CaFA and PUFAR, respectively, and increased DMI by 0.55 kg/d in SFAR. Correctedmilk yield increased by 0.44 kg/d for each percentage-unit increase in crude fat level. Plasma non-esterified fatty acid (NEFA) concentration increased with 0.04 mmol/L per percentage-unit increase. Inconclusion, the increased crude fat level increased corrected milk yield. Fat supplementation in earlylactation does not seem to cause negative effects on productive performance or metabolism, anddespite a small increase, both plasma NEFA and β-hydroxybutyrate (BHB) concentrations stayed belowthe threshold for incidence of subclinical ketosis.

A functional connection between the Microprocessor and a variant NEXT complex

Imamura, K., Garland, W., Schmid, M., et al. — Fri, 01 Nov 2024 20:36:23 +0100

In mammalian cells, primary miRNAs are cleaved at their hairpin structures by the Microprocessor complex, whose core is composed of DROSHA and DGCR8. Here, we show that 5′ flanking regions, resulting from Microprocessor cleavage, are targeted by the RNA exosome in mouse embryonic stem cells (mESCs). This is facilitated by a physical link between DGCR8 and the nuclear exosome targeting (NEXT) component ZCCHC8. Surprisingly, however, both biochemical and mutagenesis studies demonstrate that a variant NEXT complex, containing the RNA helicase MTR4 but devoid of the RNA-binding protein RBM7, is the active entity. This Microprocessor-NEXT variant also targets stem-loop-containing RNAs expressed from other genomic regions, such as enhancers. By contrast, Microprocessor does not contribute to the turnover of less structured NEXT substrates. Our results therefore demonstrate that MTR4-ZCCHC8 can link to either RBM7 or DGCR8/DROSHA to target different RNA substrates depending on their structural context.

Publications - Peer-reviewed articles

A randomized controlled trial investigating the neurocognitive effects of Lacprodan® PL-20, a phospholipid-rich milk protein concentrate, in elderly participants with age-associated memory impairment

Dietary milk-fat-globule membrane affects resistance to diarrheagenic escherichia coli in healthy adults in a randomized, placebo-controlled, double-blind study

Effects of milk-based phospholipids on cognitive performance and subjective responses to psychosocial stress

Inorganic phosphate transporter PiT2 and phosphate homeostasis

Exploiting O-GlcNAc dyshomeostasis to screen O-GlcNAc transferase intellectual disability variants

Palatinale rugae til human identifikation:

Structure and Function of the Leukocyte Integrin αMβ2

Characterization of the bispecific VHH antibody tarperprumig (ALXN1820) specific for properdin and designed for low-volume administration

Unveiling the enzymatic pathway of UMG-SP2 urethanase

Control of neonatal diarrhea in piglets with reduced antibiotic use by application of a complementary feed - a randomized controlled farm trial

Single-Molecule Multivalent Interactions Revealed by Plasmon-Enhanced Fluorescence

Shared genetic risk between anorexia nervosa and cardiovascular disease events

Structural and biochemical analysis of ligand binding in yeast Niemann–Pick type C1–related protein

Analysis of exonic deletions in a large population study provides novel insights into NRXN1 pathology

Alkoxy Substituted Brominated closo-Dodecaborates with Functionalized Aliphatic Spacers

Phosphine Oxide Indenoquinoline Derivatives

TrAnnoScope

Dynamics of IGF Signaling During the Ovulatory Peak in Women Undergoing Ovarian Stimulation

The Cryo-EM structure of human CD163 bound to haptoglobin-hemoglobin reveals molecular mechanisms of hemoglobin scavenging

Optimized Construction of a Yeast SICLOPPS Library for Unbiased In Vivo Selection of Cyclic Peptides

Resilience and Charge-Dependent Fibrillation of functional amyloid

Modeling of mRNA deadenylation rates reveal a complex relationship between mRNA deadenylation and decay

Rescuable sleep and synaptogenesis phenotypes in a Drosophila model of O-GlcNAc transferase intellectual disability

Exercise, hormesis and ageing

Non-Invasive Malaria Detection in Sub-Saharan Africa Using a DNA-Based Sensor System

DNA-directed termination of mammalian RNA polymerase II

RNA 3′end tailing safeguards cells against products of pervasive transcription termination

Increased diversity of beneficial rhizobia enhances faba bean growth

A cautionary tale for Alzheimer’s disease GWAS by proxy

Enhancing Membrane Permeability of Fluorescein-Type Chromophore Through Covalent Attachment of Chlorinated Dodecaborate

Dimethyl labeling of N-terminal amines allows unambiguous identification of protein crosslinks

Nanobodies' duo facilitates ultrasensitive serum HER-2/neu immunoassays via enhanced avidity interactions

Distinct fingerprints of tRNA-derived small non-coding RNA in animal models of neurodegeneration

Young rat microbiota extracts strongly inhibit fibrillation of α-synuclein and protect neuroblastoma cells and zebrafish against α-synuclein toxicity

m6A modification of mutant huntingtin RNA promotes the biogenesis of pathogenic huntingtin transcripts

Hydrogel bead-based isothermal detection (BEAD-ID) for assessing the activity of DNA-modifying enzymes

Hydrogel bead-based isothermal detection (BEAD-ID) for assessing the activity of DNA-modifying enzymes

Compartment-specific small non-coding RNA changes and nucleolar defects in human mesial temporal lobe epilepsy

A conserved juxtamembrane motif in plant NFR5 receptors is essential for root nodule symbiosis

Magnetoresponsive liposomes applications in nanomedicine

Extensive Population Structure Highlights an Apparent Paradox of Stasis in the Impala (Aepyceros melampus)

DiDBiT-TMT

Natural variation in root exudate composition in the genetically structured Arabidopsis thaliana in the Iberian Peninsula

Genetic liability estimated from large-scale family data improves genetic prediction, risk score profiling, and gene mapping for major depression

The effects of Zanthoxylum bungeanum extract on lipid metabolism induced by sterols

Novel metabolic role for BDNF in pancreatic β-cell insulin secretion

Generation of Functional Mouse Hippocampal Neurons

TrkA-cholinergic signaling modulates fear encoding and extinction learning in PTSD-like behavior

Fat supplement for dairy cows during early lactation – potentials, challenges, and risks – a meta-analysis

A functional connection between the Microprocessor and a variant NEXT complex

m⁶A modification of mutant huntingtin RNA promotes the biogenesis of pathogenic huntingtin transcripts