bluuebunny/crossref_metadata_2025 huggingface dataset:
Maternal diabetes influences neonatal obesity-adiposity but not in later life Offspring obesity in diabetic pregnancy
Authors: Deshpande-Joshi Sayali S. , Wagle Sonali S. , Deshmukh Madhura K. , Damle Hemant S. , Otiv Suhas R. , Phatak Sanat B. , Dhadge Smita N. , Ambardekar Shubha S. , Bhat Dattatray S. , Raut Deepa A. , Kamat Rajashree P. , Wadke Sayali G. , Kumaran Kalyanaraman , Chandak Giriraj R. , Yajnik Chittaranjan S.
Published: May 2023

Background

Based on studies in overweight-obese populations, it is tacitly assumed that maternal hyperglycemia is responsible for obesity-adiposity at birth and in later life.

Study design

Two hospital based case control studies: 1) Neonatal outcomes, 2) Later life outcomes.

Methods

We studied associations of neonatal and later life obesity-adiposity [age and sex-adjusted BMI, waist circumference, skinfolds, and body fat percent by Dual energy X-ray Absorptiometry (DXA)] in offspring of mothers with diabetes (ODM) and those of mothers without diabetes (ONDM). Exposures were parental hyperglycemia and overweight-obesity.

Results

Neonatal study included 372 non-diabetic and 816 diabetic pregnancies [74 type 1 diabetes, 102 type 2 diabetes, 640 gestational diabetes (GDM)]. Mothers with type 1 diabetes were the youngest, thinnest, and with highest HbA1c. Maternal glycemia but not BMI was associated with neonatal obesity-adiposity. Thus, neonates of mothers with type 1 diabetes had highest ponderal index, abdominal circumference, and skinfolds.

Later life study included 200 ODM (25 type 1 diabetes, 22 type 2 diabetes, 153 GDM) and 177 age, sex and socio-economic matched ONDM (2 to 26 y). Their obesity-adiposity was associated with bi-parental overweight-obesity in an additive manner, but not with parental diabetes. Offspring birth weight was also positively associated. Offspring of mothers with type 1 diabetes had the lowest and offspring of mothers with type 2 diabetes the highest obesity-adiposity.

Conclusion

Neonatal obesity-adiposity is driven by maternal glycemia while later life obesity-adiposity by bi-parental obesity. Our results provide a clear insight into pathogenesis of obesity-adiposity in the offspring.

Article Highlights

It is tacitly assumed that maternal diabetes is responsible for offspring obesity-adiposity.

We examined the determinants of obesity-adiposity in intrauterine and in later life in children born to mothers with type 1, type 2 and GDM. Paternal influence was also investigated.

Mothers with type 1 diabetes were the thinnest and most hyperglycemic. Their children were the most obese-adipose at birth but thinnest in later life. Later life obesity-adiposity was driven by bi-parental overweight-obesity, not by diabetes.

Our findings suggest that strict maternal metabolic control during pregnancy will reduce macrosomia while targeting obesogenic family environment may reduce later life offspring obesity-adiposity.
Urinary peptides predict future death
Authors: Keller Felix , Beige Joachim , Siwy Justyna , Mebazaa Alexandre , An Dewei , Mischak Harald , Schanstra Joost P. , Mokou Marika , Perco Paul , Staessen Jan A. , Vlahou Antonia , Latosinska Agnieszka
Published: May 2023

Background

There is evidence of pre-established vulnerability in individuals that increases the risk of their progression to severe disease or death, though the mechanisms that cause this are still not fully understood. Previous research has demonstrated that a urinary peptide classifier (COV50) predicts disease progression and death from SARS-CoV-2 at an early stage, indicating that the outcome prediction may be partly due to already present vulnerabilities. The aim of this study is to examine the ability of COV50 to predict future non-COVID-19-related mortality, and evaluate whether the pre-established vulnerability can be generic and explained on a molecular level by urinary peptides.

Methods

Urinary proteomic data from 9193 patients (1719 patients sampled at intensive care unit (ICU) admission and 7474 patients with other diseases (non-ICU)) were extracted from the Human Urinary Proteome Database. The previously developed COV50 classifier, a urinary proteomics biomarker panel consisting of 50 peptides, was applied to all datasets. The association of COV50 scoring with mortality was evaluated.

Results

In the ICU group, an increase in the COV50 score of one unit resulted in a 20% higher relative risk of death (adj. HR 1·2 [95% CI 1·17-1·24]). The same increase in COV50 in non-ICU patients resulted in a higher relative risk of 61% (adj. HR 1·61 [95% CI 1·47-1·76]), in line with adjusted meta-analytic HR estimate of 1·55. The most notable and significant changes associated with future fatal events were reductions of specific collagen fragments, most of collagen alpha I(I).

Conclusion

The COV50 classifier is predictive of death in the absence of SARS-CoV-2 infection, suggesting that it detects pre-existing vulnerability. Prediction is based mainly on collagen fragments, possibly reflecting disturbances in the integrity of the extracellular matrix. These data may serve as basis for proteomics guided intervention aiming towards manipulating/improving collagen turnover, thereby reducing the risk of death.
Mediators of monocyte chemotaxis and matrix remodeling are associated with the development of fibrosis in patients with COVID-19
Authors: Holton Sarah E. , Mitchem Mallorie , Pipavath Sudhakar , Morrell Eric D. , Bhatraju Pavan K. , Hamerman Jessica A. , Speake Cate , Malhotra Uma , Wurfel Mark M. , Ziegler Steven , Mikacenic Carmen
Published: May 2023

Acute respiratory distress syndrome (ARDS) has a fibroproliferative phase that may be followed by pulmonary fibrosis. This has been described in patients with COVID-19 pneumonia, but the underlying mechanisms have not been completely defined. We hypothesized that protein mediators of tissue remodeling and monocyte chemotaxis are elevated in the plasma and endotracheal aspirates of critically ill patients with COVID-19 who subsequently develop radiographic fibrosis.

We enrolled COVID-19 patients admitted to the ICU who had hypoxemic respiratory failure, were hospitalized and alive for at least 10 days, and had chest imaging done during hospitalization (n= 119). Plasma was collected within 24h of ICU admission and at 7d. In mechanically ventilated patients, endotracheal aspirates (ETA) were collected at 24h and 48-96h. Protein concentrations were measured by immunoassay. We tested for associations between protein concentrations and radiographic evidence of fibrosis using logistic regression adjusting for age, sex, and APACHE score.

We identified 39 patients (33%) with features of fibrosis. Within 24h of ICU admission, plasma proteins related to tissue remodeling (MMP-9, Amphiregulin) and monocyte chemotaxis (CCL-2/MCP-1, CCL-13/MCP-4) were associated with the subsequent development of fibrosis whereas markers of inflammation (IL-6, TNF-α) were not. After 1 week, plasma MMP-9 increased in patients without fibrosis. In ETAs, only CCL-2/MCP-1 was associated with fibrosis at the later timepoint.

This cohort study identifies proteins of tissue remodeling and monocyte recruitment that may identify early fibrotic remodeling following COVID-19. Measuring changes in these proteins over time may allow for early detection of fibrosis in patients with COVID-19.
Neighborhood air pollution is negatively associated with neurocognitive maturation in early adolescence
Authors: Kardan Omid , Sereeyothin Chacriya , Schertz Kathryn E. , Angstadt Mike , Weigard Alexander S. , Berman Marc G. , Heitzeg Mary M. , Rosenberg Monica D.
Published: May 2023

The ability to maintain focus and process task-relevant information continues developing during adolescence, but the specific physical environmental factors that influence this development remain poorly characterized. One candidate factor is air pollution. Evidence suggests that small particulate matter and NO2 concentrations in the air may negatively impact cognitive development in childhood. We assessed the relationship between neighborhood air pollution and the changes in performance on then-back task, a test of attention and working memory, in the Adolescent Brain Cognitive Development (ABCD) Study’s baseline (ages 9-10) and two-year-follow-up releases (Y2, ages 11-12; n = 5,256). In thebehavioraldomain, multiple linear regression showed that developmental change in n-back task performance was negatively associated with neighborhood air pollution (β = -.044, t = -3.11, p = .002), adjusted for covariates capturing baseline cognitive performance of the child, their parental income and education, family conflicts, and their neighborhood’s population density, crime rate, perceived safety, and Area Deprivation Index (ADI). The strength of the adjusted association for air pollution was similar to parental income, family conflict, and neighborhood ADI. In theneuroimagingdomain, we evaluated a previously published youthcognitivecomposite Connectome-based Predictive Model (ccCPM), and again found that decreased developmental change in the strength of the ccCPM from pre-to early adolescence was associated with neighborhood air pollution (β = -.110, t = -2.69, p = .007), adjusted for the covariates mentioned above and head motion. Finally, we found that the developmental change in ccCPM strength was predictive of the developmental change in n-back performance (r= .157, p < .001), and there was an indirect-only mediation where the effect of air pollution on change in n-back performance was mediated by the change in the ccCPM strength (βindirecteffect= -.013, p = .029). In conclusion, neighborhood air pollution is associated with lags in the maturation of youth cognitive performance and decreased strengthening of the brain networks supporting cognitive abilities over time.
Assessment of valvular function in over 47,000 people using deep learning-based flow measurements
Authors: Kany Shinwan , Rämö Joel T. , Hou Cody , Jurgens Sean J. , Nauffal Victor , Cunningham Jon , Lau Emily S. , Butte Atul J. , Ho Jennifer E. , Olgin Jeffrey E. , Elmariah Sammy , Lindsay Mark E. , Ellinor Patrick T. , Pirruccello James P.
Published: May 2023

Valvular heart disease is associated with a high global burden of disease. Even mild aortic stenosis confers increased morbidity and mortality, prompting interest in understanding normal variation in valvular function at scale.

We developed a deep learning model to study velocity-encoded magnetic resonance imaging in 47,223 UK Biobank participants. We calculated eight traits, including peak velocity, mean gradient, aortic valve area, forward stroke volume, mitral and aortic regurgitant volume, greatest average velocity, and ascending aortic diameter. We then computed sex-stratified reference ranges for these phenotypes in up to 31,909 healthy individuals. In healthy individuals, we found an annual decrement of 0.03cm2in the aortic valve area. Participants with mitral valve prolapse had a 1 standard deviation [SD] higher mitral regurgitant volume (P=9.6 × 10−12), and those with aortic stenosis had a 4.5 SD-higher mean gradient (P=1.5 × 10−431), validating the derived phenotypes’ associations with clinical disease. Greater levels of ApoB, triglycerides, and Lp(a) assayed nearly 10 years prior to imaging were associated with higher gradients across the aortic valve. Metabolomic profiles revealed that increased glycoprotein acetyls were also associated with an increased aortic valve mean gradient (0.92 SD, P=2.1 x 10−22). Finally, velocity-derived phenotypes were risk markers for aortic and mitral valve surgery even at thresholds below what is considered relevant disease currently.

Using machine learning to quantify the rich phenotypic data of the UK Biobank, we report the largest assessment of valvular function and cardiovascular disease in the general population.
Y-chromosomal microsatellites and expansion from Africa
Authors: Cenac Zarus
Published: May 2023

Research has suggested that Africa is the continent from which modern humans dispersed around the world. Some diversities are thought to trace this expansion; they decline as distance from Africa accumulates, and they suggest that the expansion originated within a range of locations in Africa. Previously, a decline was found regarding Y-chromosomal microsatellite heterozygosity. However, this diversity has been noted to suggest a non-African origin and, consequently, not look like it indicates expansion from Africa. Declines have appeared in other variables which are derived from Y-chromosomal microsatellites. These variables include effective population size, expansion time, and time to the most recent common ancestor. The present research inferred if these variables, and some Y-chromosomal diversities, indicate expansion from Africa. Variables which indicate the expansion could help with identifying which area of Africa the expansion started in. This research used variables previously derived from Y-chromosomal microsatellites. These variables were for populations from across the world (effective population size, expansion time, and time to the most recent common ancestor) or only in Africa (haplotype and repeat unit diversities). Regarding populations across the world, San were very high in each variable when considering distance from (southern) Africa. Without San, effective population size and time to the most recent common ancestor did not squarely indicate an African origin, and expansion time placed the origin wholly outside of Africa. When using San, each area was only in Africa. Amongst African populations, whilst both haplotype diversity and repeat unit diversity showed a geographical pattern of declining, only haplotype diversity suggested a completely African area of origin. A southern African origin was broadly indicated when a general estimate of the origin was produced from Y-chromosomal haplotype diversity alongside some other genetic diversities.
Geographic pair-matching in large-scale cluster randomized trials
Authors: Arnold Benjamin F. , Rerolle Francois , Tedijanto Christine , Njenga Sammy M. , Rahman Mahbubur , Ercumen Ayse , Mertens Andrew , Pickering Amy , Lin Audrie , Arnold Charles D. , Das Kishor , Stewart Christine P. , Null Clair , Luby Stephen P. , Colford John M. , Hubbard Alan E. , Benjamin-Chung Jade
Published: May 2023

Custer randomized trials are often used to study large-scale public health interventions. In large trials, even small improvements in statistical efficiency can have profound impacts on the required sample size and cost. Pair matched randomization is one strategy with potential to increase trial efficiency, but to our knowledge there have been no empirical evaluations of pair-matching in large-scale, epidemiologic field trials. Location integrates many socio-demographic and environmental characteristics into a single feature. Here, we show that geographic pair-matching leads to substantial gains in statistical efficiency for 14 child health outcomes that span growth, development, and infectious disease through a re-analysis of two large-scale trials of nutritional and environmental interventions in Bangladesh and Kenya. We estimate relative efficiencies ≥ 1.1 for all outcomes assessed and relative efficiencies regularly exceed 2.0, meaning an unmatched trial would have needed to enroll at least twice as many clusters to achieve the same level of precision as the geographically pair-matched design. We also show that geographically pair-matched designs enable estimation of fine-scale, spatially varying effect heterogeneity under minimal assumptions. Our results demonstrate broad, substantial benefits of geographic pair-matching in large-scale, cluster randomized trials.
Repeated Omicron exposures override ancestral SARS-CoV-2 immune imprinting
Authors: Yisimayi Ayijiang , Song Weiliang , Wang Jing , Jian Fanchong , Yu Yuanling , Chen Xiaosu , Xu Yanli , Yang Sijie , Niu Xiao , Xiao Tianhe , Wang Jing , Zhao Lijuan , Sun Haiyan , An Ran , Zhang Na , Wang Yao , Wang Peng , Yu Lingling , Lv Zhe , Gu Qingqing , Shao Fei , Jin Ronghua , Shen Zhongyang , Xie Xiaoliang Sunney , Wang Youchun , Cao Yunlong
Published: May 2023

The continuous emergence of highly immune evasive SARS-CoV-2 variants, like XBB.1.51,2and XBB.1.163,4, highlights the need to update COVID-19 vaccine compositions. However, immune imprinting induced by wildtype (WT)-based vaccination would compromise the antibody response to Omicron-based boosters5-9. Vaccination strategies that can counter immune imprinting are critically needed. In this study, we investigated the degree and dynamics of immune imprinting in mouse models and human cohorts, especially focusing on the role of repeated Omicron stimulation. Our results show that in mice, the efficacy of single Omicron-boosting is heavily limited by immune imprinting, especially when using variants antigenically distinct from WT, like XBB, while the concerning situation could be largely mitigated by a second Omicron booster. Similarly, in humans, we found that repeated Omicron infections could also alleviate WT-vaccination-induced immune imprinting and generate high neutralizing titers against XBB.1.5 and XBB.1.16 in both plasma and nasal mucosa. By isolating 781 RBD-targeting mAbs from repeated Omicron infection cohorts, we revealed that double Omicron exposure alleviates immune imprinting by generating a large proportion of highly matured and potent Omicron-specific antibodies. Importantly, epitope characterization using deep mutational scanning (DMS) showed that these Omicron-specific antibodies target distinct RBD epitopes compared to WT-induced antibodies, and the bias towards non-neutralizing epitopes observed in single Omicron exposures due to imprinting was largely restored after repeated Omicron stimulation, together leading to a substantial neutralizing epitope shift. Based on the DMS profiles, we identified evolution hotspots of XBB.1.5 RBD and demonstrated the combinations of these mutations could further boost XBB.1.5’s immune-evasion capability while maintaining high ACE2 binding affinity. Our findings suggest the WT component should be abandoned when updating COVID-19 vaccine antigen compositions to XBB lineages, and those who haven’t been exposed to Omicron yet should receive two updated vaccine boosters.
An expanded palette of ATP sensors for subcellular and multiplexed imaging
Authors: Sahan Ayse Z. , Youngblood Eventine , Das Siddharth , Schmitt Danielle L. , Zhang Jin
Published: May 2023

Genetically encoded fluorescent biosensors that detect changes in ATP levels in live cells have enabled the discovery of novel roles for ATP in cellular processes and signaling. Many of these available ATP biosensors have a limited dynamic range, or have ATP affinities that are not suitable for sensing the physiological concentrations of ATP in mammalian cells. To address these limitations, we developed a FRET-based ATP biosensor with enhanced dynamic range and signal to noise ratio, eATeam. Using eATeam, we uncovered distinct spatiotemporal dynamics of ATP changes upon inhibition of cellular energy production. We also developed dimerization-dependent GFP and RFP-based ATP biosensors with enhanced dynamic ranges compared to the current standard in the field. Using the single-color ATP biosensors, we visualized the complex interplay between AMPK activity, ATP, lactate, and calcium by multiplexed imaging in single cells. This palette of ATP sensors expands the toolbox for interrogating subcellular ATP regulation and metabolic signaling in living cells.
Regulation of lipolysis by 14-3-3 proteins on human adipocyte lipid droplets
Authors: Yang Qin , Loureiro Zinger Yang , Desai Anand , DeSouza Tiffany , Li Kaida , Wang Hui , Nicoloro Sarah M , Solivan-Rivera Javier , Corvera Silvia
Published: May 2023

Adipocyte lipid droplets (LDs) play a crucial role in systemic lipid metabolism by storing and releasing lipids to meet the organism’s energy needs. Hormonal signals such as catecholamines and insulin act on adipocyte LDs, and impaired responsiveness to these signals can lead to uncontrolled lipolysis, lipotoxicity, and a higher risk of metabolic diseases. To investigate the mechanisms that control LD function in human adipocytes, we employed techniques to obtain mesenchymal progenitor cells on a large scale and applied proximity labeling mediated by enhanced ascorbate peroxidase (APEX2) to identify the interactome of PLIN1 in differentiated adipocytes. We identified 70 proteins that interact specifically with PLIN1, including PNPLA2 and LIPE, which are the primary effectors of regulated triglyceride hydrolysis, and four members of the 14-3-3 protein family (YWHAB, YWHAE, YWHAZ, and YWHAG), which are known to regulate diverse signaling pathways. Functional studies showed that YWHAB is required for maximum cAMP-stimulated lipolysis and helps to mitigate the anti-lipolytic effects of insulin. These findings reveal new regulatory mechanisms that control lipolysis in human metabolism.

SIGNIFICANCE STATEMENT

Lipid droplets are ubiquitous cytoplasmic organelles that store metabolic energy and play a key role in cellular lipid metabolism (1). Adipocyte LDs play an additional, crucial role, as they supply the energy needs of the whole body through hormonally regulated triglyceride synthesis, storage, and release. The mechanisms by which adipocyte lipid droplets release lipids for systemic use has been mostly studied in mouse models and cell lines. To understand how lipid mobilization is controlled in human adipocytes, we used proximity labeling to identify proteins that interact with PLIN1, a major component of the lipid droplet, in adipocytes generated from primary human progenitor cells. Our study catalogues the interactome of human PLIN1 and identifies previously unrecognized potential mechanism for control of human adipocyte lipolysis through 14-3-3 proteins.
Glycogen phosphorylase inhibition alongside taxol chemotherapy synergistically elicits ferroptotic cell death in clear cell ovarian and kidney cancers
Authors: Khan Tashbib , Kryza Thomas , He Yaowu , Gunter Jennifer H , Gough Madeline , Snell Cameron , Hooper John D.
Published: May 2023

Background

Clear cell carcinomas (CCCs) are a distinct histopathological subtype defined by a clear cytoplasm comprised of glycogen and lipids and characterised by poor prognosis and widespread chemoresistance. In the present work we investigate glycogen metabolism as a targetable modality for these cancers.

Methods and Results

Adopting the indole carboxamide site pan-glycogen phosphorylase inhibitor CP91149 against clear cell ovarian and renal cancer cell line models, we note antiproliferative and antimigratory effects, as well as energetic stress reflected by reduced ATP pools and increased superoxide-derived reactive oxygen species. Following this, using the agent alongside standard of care chemotherapies for clear cell ovarian (ccOC) and renal cell carcinoma (ccRCC), we note specific synergy with microtubule disrupting chemotherapy paclitaxel, a phenomenon retained in ccOC lines made stably resistant to paclitaxel. Rescue experiments, as well as phenotypic assays suggest that combination-treated cells undergo ferroptotic cell death. We postulate this synergistic efficacy to arise from subjecting the already hypersensitive clear cell cancers to the mitochondrial stress elicited by taxol chemotherapy alongside the oxidative stress augured by glycogen phosphorylase inhibition.

Conclusions

Given that CCCs are widely chemoresistant, the present work potentially presents a novel therapeutic avenue for this shared histotype.
KairoSight-3.0: A Validated Optical Mapping Software to Characterize Cardiac Electrophysiology, Excitation-Contraction Coupling, and Alternans
Authors: Haq Kazi T , Roberts Anysja , Berk Fiona , Allen Samuel , Swift Luther M , Posnack Nikki Gillum
Published: May 2023

Background

Cardiac optical mapping is an imaging technique that measures fluorescent signals across a cardiac preparation. Dual optical mapping of voltage-sensitive and calcium-sensitive probes allow for simultaneous recordings of cardiac action potentials and intracellular calcium transients with high spatiotemporal resolution. The analysis of these complex optical datasets is both time intensive and technically challenging; as such, we have developed a software package for semi-automated image processing and analysis. Herein, we report an updated version of our software package (KairoSight-3.0) with features to enhance characterization of cardiac parameters using optical signals.

Methods

To test software validity and applicability, we used Langendorff-perfused heart preparations to record transmembrane voltage and intracellular calcium signals from the epicardial surface. Isolated hearts from guinea pigs and rats were loaded with a potentiometric dye (RH237) and/or calcium indicator dye (Rhod-2AM) and fluorescent signals were acquired. We used Python 3.8.5 programming language to develop theKairoSight-3.0software. Cardiac maps were validated with a user-specified manual mapping approach.

Results

Manual maps of action potential duration (30 or 80% repolarization), calcium transient duration (30 or 80% reuptake), action potential and calcium transient alternans were constituted to validate the accuracy of software-generated maps. Manual and software maps had high accuracy, with >97% of manual and software values falling within 10 ms of each other and >75% within 5 ms for action potential duration and calcium transient duration measurements (n=1000-2000 pixels). Further, our software package includes additional cardiac metric measurement tools to analyze signal-to-noise ratio, conduction velocity, action potential and calcium transient alternans, and action potential-calcium transient coupling time to produce physiologically meaningful optical maps.

Conclusions

KairoSight-3.0has enhanced capabilities to perform measurements of cardiac electrophysiology, calcium handling, and the excitation-contraction coupling with satisfactory accuracy.

Graphical Abstract Demonstrating Experimental and Data Analysis Workflow

Created withBiorender.com
Unveiling the Catalytic Mechanism of GTP Hydrolysis in Microtubules
Authors: Beckett Daniel , Voth Gregory A.
Published: May 2023

Microtubules (MTs) are large cytoskeletal polymers, composed of αβ-tubulin heterodimers, capable of stochastically converting from polymerizing to depolymerizing states and vice-versa. Depolymerization is coupled with hydrolysis of GTP within β-tubulin. Hydrolysis is favored in the MT lattice compared to free heterodimer with an experimentally observed rate increase of 500 to 700 fold, corresponding to an energetic barrier lowering of 3.8 to 4.0 kcal/mol. Mutagenesis studies have implicated α-tubulin residues, α:E254 and α:D251, as catalytic residues completing the β-tubulin active site of the lower heterodimer in the MT lattice. The mechanism for GTP hydrolysis in the free heterodimer, however, is not understood. Additionally, there has been debate concerning whether the GTP-state lattice is expanded or compacted relative to the GDP-state and whether a “compacted” GDP-state lattice is required for hydrolysis. In this work, extensive QM/MM simulations with transition-tempered metadynamics free energy sampling of compacted and expanded inter-dimer complexes, as well as free heterodimer, have been carried out to provide clear insight into the GTP hydrolysis mechanism. α:E254 was found to be the catalytic residue in a compacted lattice, while in the expanded lattice disruption of a key salt bridge interaction renders α:E254 less effective. The simulations reveal a barrier decrease of 3.8 ± 0.5 kcal/mol for the compacted lattice compared to free heterodimer, in good agreement with experimental kinetic measurements. Additionally, the expanded lattice barrier was found to be 6.3 ± 0.5 kcal/mol higher than compacted, demonstrating that GTP hydrolysis is variable with lattice state and slower at the MT tip.

Significance Statement

Microtubules (MTs) are large and dynamic components of the eukaryotic cytoskeleton with the ability to stochastically convert from a polymerizing to a depolymerizing state and vice-versa. Depolymerization is coupled to the hydrolysis of guanosine-5’-triphosphate (GTP), which is orders of magnitude faster in the MT lattice than in free tubulin heterodimers. Our results computationally ascertain the catalytic residue contacts in the MT lattice that accelerate GTP hydrolysis compared to the free heterodimer as well as confirm that a compacted MT lattice is necessary for hydrolysis while a more expanded lattice is unable to form the necessary contacts and thereby hydrolyze GTP.
Improving AlphaFold2-based Protein Tertiary Structure Prediction with MULTICOM in CASP15
Authors: Liu Jian , Guo Zhiye , Wu Tianqi , Roy Raj S. , Chen Chen , Cheng Jianlin
Published: May 2023

Since CASP14, AlphaFold2 has become the standard method for protein tertiary structure prediction. One remaining challenge in the field is to further improve the accuracy of AlphaFold2-based protein structure prediction. To address this challenge, we developed a new version of the MULTICOM system to sample diverse multiple sequence alignments (MSAs) and structural templates to improve the input for AlphaFold2 to generate structural models. The models are then ranked by both the pairwise model similarity and AlphaFold2 self-reported model quality score. The top ranked models are further refined by a novel structure alignment-based refinement method powered by Foldseek. Moreover, for a monomer target that is a subunit of a protein assembly (complex), MULTICOM integrates tertiary and quaternary structure prediction together to account for tertiary structural changes induced by protein-protein interaction in the assembly. The MULTICOM system participated in the tertiary structure prediction in the 15thCritical Assessment of Techniques for Protein Structure Prediction (CASP15) in 2022 as server and human predictors. Our best server predictor (MULTICOM_refine) ranked 3rdamong 47 CASP15 server predictors and our best human predictor (MULTICOM) ranked 7thamong all 132 human and server predictors. The average GDT-TS score and TM-score of the first structural models that MULTICOM_refine predicted for 94 CASP15 domains are ∼0.80 and ∼0.92, 9.6% and 8.2% and higher than ∼0.73 and 0.85 of the standard AlphaFold2 predictor respectively. The results demonstrate that our approach can significantly improve the accuracy of the AlphaFold2-based protein tertiary structure prediction. The source code of MULTICOM is available at:https://github.com/BioinfoMachineLearning/MULTICOM3.
Lysine-independent ubiquitination and degradation of REV-ERBα involves a bi-functional degradation control sequence at its N-terminus
Authors: Suen Ting-Chung , DeBruyne Jason P.
Published: May 2023

REV-ERBα and REV-ERBβ proteins play crucial roles in linking the circadian system to overt daily rhythms in mammalian physiology and behavior. In most tissues, REV-ERBα protein robustly cycles such that it is detected only within a tight interval of 4-6 hours each day, suggesting both its synthesis and degradation are tightly controlled. Several ubiquitin ligases are known to drive REV-ERBα degradation, but how they interact with REV-ERBα and which lysine residues they ubiquitinate to promote degradation are unknown. In this study, we attempted to identify both ubiquitin-ligase-binding and ubiquitination sites within REV-ERBα required for its degradation. Surprisingly, mutating all lysine residues, the common sites for ubiquitin conjugation, in REV-ERBα to arginines (K20R), did very little to impair its degradation in cells. K20R were degraded much faster by co-expression of two E3 ligases, SIAH2 or SPSB4, suggesting possible N-terminal ubiquitination. To explore this, we examined if small deletions at the N-terminus of REV-ERBα would alter its degradation. Interestingly, deletion of amino acid (AA) residues 2 to 9 (delAA2-9) clearly resulted in a less stable REV-ERBα. We found that it was the length (i.e. 8 AA), and not the specific sequence, that confers stability in this region. Simultaneously, we also mapped the interaction site of the E3 ligase SPSB4 to this same region, specifically requiring AA4-9 of REV-ERBα. Thus, the first 9 AA of REV-ERBα has two opposing roles in regulating REV-ERBα turnover. Further, deleting eight additional AAs (delAA2-17) from the N-terminus strongly prevents REV-ERBα degradation. Combined, these results suggest that complex interactions within the first 25AAs potentially act as an endogenous ‘switch’ that allows REV-ERBα to exist in a stabilized conformation in order to accumulate at one time of day, but then rapidly shifts to a destabilized form, to enhance its removal at the end of its daily cycle.
Functional genomic analysis of non-canonical DNA regulatory elements of the aryl hydrocarbon receptor
Authors: Patel Tajhal D , Nakka Manjula , Grimm Sandra L , Coarfa Cristian , Gorelick Daniel A
Published: May 2023

The aryl hydrocarbon receptor (AHR) is a ligand-dependent transcription factor that binds DNA and regulates genes in response to halogenated and polycyclic aromatic hydrocarbons. AHR also regulates the development and function of the liver and the immune system. In the canonical pathway, AHR binds a consensus DNA sequence, termed the xenobiotic response element (XRE), recruits protein coregulators, and regulates target gene expression. Emerging evidence suggests that AHR may regulate gene expression via an additional pathway, by binding to a non-consensus DNA sequence termed the non-consensus XRE (NC-XRE). The prevalence of NC-XRE motifs in the genome is not known. Studies using chromatin immunoprecipitation and reporter genes provide indirect evidence of AHR-NC-XRE interactions, but direct evidence for an AHR-NCXRE interaction that regulates transcription in a natural genomic context is lacking. Here, we analyzed AHR binding to NC-XRE DNA on a genome-wide scale in mouse liver. We integrated ChIP-seq and RNA-seq data and identified putative AHR target genes with NC-XRE motifs in regulatory regions. We also performed functional genomics at a single locus, the mouseSerpine1gene. Deleting NC-XRE motifs from theSerpine1promoter reduced the upregulation ofSerpine1by TCDD, an AHR ligand. We conclude that AHR upregulatesSerpine1via NC-XRE DNA. NC-XRE motifs are prevalent throughout regions of the genome where AHR binds. Taken together, our results suggest that AHR regulates genes via NC-XRE motifs. Our results will also improve our ability to identify AHR target genes and their physiologic relevance.
GPCR binding and JNK3 activation by arrestin-3 have different structural requirements
Authors: Zheng Chen , Weinstein Liana D. , Nguyen Kevin K. , Grewal Abhijeet , Gurevich Eugenia V. , Gurevich Vsevolod V.
Published: May 2023

Arrestins bind active phosphorylated G protein-coupled receptors (GPCRs). Among the four mammalian subtypes, only arrestin-3 facilitates the activation of JNK3 in cells. In available structures, Lys-295 in the lariat loop of arrestin-3 and its homologue Lys-294 in arrestin-2 directly interact with the activator-attached phosphates. We compared the role of arrestin-3 conformational equilibrium and of Lys-295 in GPCR binding and JNK3 activation. Several mutants with enhanced ability to bind GPCRs showed much lower activity towards JNK3, whereas a mutant that does not bind GPCRs was more active. Subcellular distribution of mutants did not correlate with GPCR recruitment or JNK3 activation. Charge neutralization and reversal mutations of Lys-295 differentially affected receptor binding on different backgrounds, but had virtually no effect on JNK3 activation. Thus, GPCR binding and arrestin-3-assisted JNK3 activation have distinct structural requirements, suggesting that facilitation of JNK3 activation is the function of arrestin-3 that is not bound to a GPCR.
The genetic code assembles via division and fusion, basic cellular events
Authors: Yarus Michael
Published: May 2023

Standard Genetic Code (SGC) evolution is quantitatively modeled in computed ‘worlds’ containing up to 2000 independent coding ‘environments’. Environments can host multiple codes that may fuse or divide, with division yielding identical descendants. Code division may be selected - sophisticated gene products could be required for orderly separation. Several unforeseen results emerge: more rapid evolution requires unselective code division, rather than its selective form. Combining selective and unselective code division, with/without code fusion, with/without independent environmental coding tables and with/without wobble defines 25= 32 possible pathways for SGC evolution. These 32 possible histories are compared, particularly, for speed and accuracy. Pathways differ greatly; for example, ≈ 300-fold different in time to evolve SGC-like codes. Eight of 32 pathways, employing code division, are quickest. Four of these eight, that combine fusion and division, also unite speed and accuracy. The two precise, swiftest paths, thus the most likely routes to the SGC, are similar, differing only in fusion with independent environmental codes. Code division instead of fusion with unrelated codes implies that independent codes can be dispensable. Instead, a single ancestral code that divides and fuses can initiate fully encoded peptide biosynthesis. Division and fusion create a ‘crescendo of competent coding’, facilitating search for the SGC, and also assist advent of otherwise disfavored wobble coding. Code fusion readily unites multiple codon assignment mechanisms. But via code division and fusion, the SGC is shown to emerge from a single primary origin, via familiar cellular events.
Automatic cell type harmonization and integration across Human Cell Atlas datasets
Authors: Xu Chuan , Prete Martin , Webb Simone , Jardine Laura , Stewart Benjamin J. , Hoo Regina , He Peng , Meyer Kerstin , Teichmann Sarah A.
Published: May 2023

Harmonizing cell types across the single-cell community and assembling them into a common framework is central to building a standardized Human Cell Atlas. Here we present CellHint, a predictive clustering tree-based tool to resolve cell type differences in annotation resolution and technical biases across datasets. CellHint accurately quantifies cell-cell transcriptomic similarities and places cell types into a relationship graph that hierarchically defines shared and unique cell subtypes. Application to multiple immune datasets recapitulates expert-curated annotations. CellHint also reveals underexplored relationships between healthy and diseased lung cell states in eight diseases. Furthermore, we present a workflow for fast cross-dataset integration guided by harmonized cell types and cell hierarchy, which uncovers underappreciated cell types in adult human hippocampus. Finally, we apply CellHint to 12 tissues from 38 datasets, providing a deeply curated cross-tissue database with ∼3.7 million cells and various machine learning models for automatic cell annotation across human tissues.
Beyond Alternans: Detection of Higher-Order Periodicity in Ex-Vivo Human Ventricles Before Induction of Ventricular Fibrillation
Authors: Iravanian Shahriar , Uzelac Ilija , Shah Anand D , Toye Mikael J , Lloyd Michael S. , Burke Michael A. , Daneshmand Mani A , Attia Tamer S , Vega J David , Merchant Faisal M. , Cherry Elizabeth M , Bhatia Neal K. , Fenton Flavio H.
Published: May 2023

Background

Repolarization alternans, defined as period-2 oscillation in the repolarization phase of the action potentials, is one of the cornerstones of cardiac electrophysiology as it provides a mechanistic link between cellular dynamics and ventricular fibrillation (VF). Theoretically, higher-order periodicities (e.g., period-4, period-8,…) are expected but have very limited experimental evidence.

Methods

We studied explanted human hearts, obtained from the recipients of heart transplantation at the time of surgery, using optical mapping technique with transmembrane voltage-sensitive fluorescent dyes. The hearts were stimulated at an increasing rate until VF was induced. The signals recorded from the right ventricle endocardial surface just before the induction of VF and in the presence of 1:1 conduction were processed using the Principal Component Analysis and a combinatorial algorithm to detect and quantify higher-order dynamics.

Results

A prominent and statistically significant 1:4 peak (corresponding to period-4 dynamics) was seen in three of the six studied hearts. Local analysis revealed the spatiotemporal distribution of higher-order periods. Period-4 was localized to temporally stable islands. Higher-order oscillations (period-5, 6, and 8) were transient and primarily occurred in arcs parallel to the activation isochrones.

Discussion

We present evidence of higher-order periodicities and the co-existence of such regions with stable non-chaotic areas in ex-vivo human hearts before VF induction. This result is consistent with the period-doubling route to chaos as a possible mechanism of VF initiation, which complements the concordant to discordant alternans mechanism. The presence of higher-order regions may act as niduses of instability that can degenerate into chaotic fibrillation.
Pannexin 1 modulates angiogenic activities of human endothelial colony-forming cells through IGF-1 mechanism and is a marker of senescence
Authors: Tien Ting-Yi , Wu Yih-Jer , Su Cheng-Huang , Hsieh Chin-Ling , Wang Bo-Jeng , Lee Yi-Nan , Su Yeu , Yeh Hung-I
Published: May 2023

BACKGROUND

We examined the role of pannexins in human endothelial progenitor cell (EPC) senescence.

METHODS

Young and replication-induced senescent endothelial colony-forming cells (ECFCs) derived from human circulating EPCs were used to examine cellular activities and senescence-associated indicators after transfection of siRNA specific to Panx1 or lentivirus-mediated Panx1 overexpression. Hindlimb ischemia mice were used asin vivoangiogenesis model. Protein and phospho-kinase arrays were used to determine underlying mechanisms.

RESULTS

Panx1 was the predominant pannexin isoform in human ECFCs and up-regulated in both replication-induced senescent ECFCs and circulating EPCs from aged mice and humans. Cellular activities of the young ECFCs were enhanced by Panx1 down-regulation, but attenuated by its up-regulation. In addition, reduction of Panx1 in the senescent ECFCs could rejuvenate cellular activities with reduced senescence-associated indicators, including senescence-associated β-galactosidase activity, p16INK4a, p21, acetyl-p53, and phospho-Histone H2A.X. In mouse ischemic hindlimbs injected senescent ECFCs, blood perfusion ratio, salvaged limb outcome, and capillary density were all improved by Panx1 knockdown. Insulin-like growth factor 1 (IGF-1) was significantly increased in the supernatant from senescent ECFCs after Panx1 knockdown. The enhanced activities and paracrine effects of Panx1 knockdown senescent ECFCs were completely inhibited by anti-IGF-1 antibodies. FAK, ERK and STAT3 were activated in senescent ECFCs with Panx1 knockdown, in which the intracellular calcium level was reduced, and the activation was inhibited by supplemented calcium. The increased IGF-1 in Panx1-knockdown ECFCs was abrogated respectively by inhibitors of FAK (PF562271), ERK (U0126), and STAT3 (NSC74859), and supplemented calcium.

CONCLUSIONS

Panx1 expression is up-regulated in human ECFCs/EPCs with replication-induced senescence and during aging. Angiogenic potential of senescent ECFCs is improved by Panx1 reduction through increased IGF-1 production via activation of FAK-ERK axis following calcium influx reduction. Our findings provide new strategies to evaluate EPC activities and rejuvenate senescent EPCs for therapeutic angiogenesis.
The endoplasmic reticulum stress sensor IRE1 regulates collagen secretion through the enforcement of the proteostasis factor P4HB/PDIA1 contributing to liver damage and fibrosis
Authors: Hazari Younis , Urra Hery , Garcia Lopez Valeria A. , Diaz Javier , Tamburini Giovanni , Milani Mateus , Pihan Philippe , Durand Sylvere , Aprahamia Fanny , Baxter Reese , Huang Menghao , Dong X Charlie , Vihinen Helena , Batista-Gonzalez Ana , Godoy Patricio , Criollo Alfredo , Ratziu Vlad , Foufelle Fabienne , Hengstler Jan G. , Jokitalo Eija , Bailly-maitre Beatrice , Maiers Jessica L , Plate Lars , Kroemer Guido , Hetz Claudio
Published: May 2023

Collagen is one the most abundant proteins and the main cargo of the secretory pathway, contributing to hepatic fibrosis and cirrhosis due to excessive deposition of extracellular matrix. Here we investigated the possible contribution of the unfolded protein response, the main adaptive pathway that monitors and adjusts the protein production capacity at the endoplasmic reticulum, to collagen biogenesis and liver disease. Genetic ablation of the ER stress sensor IRE1 reduced liver damage and diminished collagen deposition in models of liver fibrosis triggered by carbon tetrachloride (CCl4) administration or by high fat diet. Proteomic and transcriptomic profiling identified the prolyl 4-hydroxylase (P4HB, also known as PDIA1), which is known to be critical for collagen maturation, as a major IRE1-induced gene. Cell culture studies demonstrated that IRE1 deficiency results in collagen retention at the ER and altered secretion, a phenotype rescued by P4HB overexpression. Taken together, our results collectively establish a role of the IRE1/P4HB axis in the regulation of collagen production and its significance in the pathogenesis of various disease states.
A unique C-terminal domain contributes to the molecular function of restorer-of-fertility proteins in plant mitochondria
Authors: Huynh Sang Dang , Melonek Joanna , Colas des Francs-Small Catherine , Bond Charles S. , Small Ian
Published: May 2023

Restorer-of-fertility(Rf) genes have practical applications in hybrid seed production as a means to control self-pollination. They encode pentatricopeptide repeat (PPR) proteins that are targeted to mitochondria where they specifically bind to transcripts that induce cytoplasmic male sterility and repress their expression. In searching for a molecular signature unique to this class of proteins, we found that a majority of known Rf proteins have a unique domain, which we called RfCTD (Restorer-of-fertility C-terminal domain), and its presence correlates with the ability to induce cleavage of the mitochondrial RNA target. We constructed a sequence profile that can quickly and accurately identify RfCTD sequences in plant genomes or transcriptomes. We screened 219 angiosperm genomes from 123 genera and found that each diploid genome encodes, on average, 25 Rf-like (RFL) proteins, of which approximately 55% contain the C-terminal signature domain. This screen also revealed considerable variation in RFL gene numbers across flowering plants. We observed that plant genera with bisexual flowers have significantly higher numbers of RFL genes compared to those with unisexual flowers, consistent with a role of these proteins in restoration of male fertility.

Finally, we show that removing the RfCTD from the RFL protein RNA PROCESSING FACTOR 2-nad6prevented cleavage of its RNA target, thenad6transcript, inArabidopsis thalianamitochondria. This research provides a simple way of identifying putativeRfcandidates in genome sequences, new insights into the molecular mode of action of Rf proteins in plant mitochondria and expands our understanding of the evolution of fertility restoration in flowering plants.