Biology subjects

Xiao, S.

Publications and source records attributed to Xiao, S..

At least 55 records · Page 3Linked to original sources

bioRxiv2025

Accurate Characterization of the Allosteric Energy Landscapes, Binding Hotspots and Long-Range Communications for KRAS Complexes with Effector Proteins : Integrative Approach Using Microsecond Molecular Dynamics, Deep Mutational Scanning of Binding Energetics and Allosteric Network Modeling

KRAS is a pivotal oncoprotein that regulates cell proliferation and survival through interactions with downstream effectors such as RAF1. Oncogenic mutations in KRAS, including G12V, G13D, and Q61R, drive constitutive activation and hyperactivation of signaling pathways, contributing to cancer progression. Despite significant advances in understanding KRAS biology, the structural and dynamic mechanisms of KRAS binding and allostery by which oncogenic mutations enhance KRAS-RAF1 binding and signaling remain incompletely understood. In this study, we employ microsecond molecular dynamics simulations, Markov State Modeling, mutational scanning and binding free energy calculations together with dynamic network modeling to elucidate the effect of KRAS mutations and characterize the thermodynamic and allosteric drivers and hotspots of KRAS binding and oncogenic activation. Our simulations revealed that oncogenic mutations stabilize the open active conformation of KRAS by differentially modulating the flexibility of the switch I and switch II regions, thereby enhancing RAF1 binding affinity. The G12V mutation rigidifies both switch I and switch II, locking KRAS in a stable, active state. In contrast, the G13D mutation moderately reduces switch I flexibility while increasing switch II dynamics, restoring a balance between stability and flexibility. The Q61R mutation induces a more complex conformational landscape, characterized by the increased switch II flexibility and expansion of functional macrostates, which promotes prolonged RAF1 binding and signaling. Mutational scanning of KRAS-RAF1 complexes identified key binding affinity hotspots, including Y40, E37, D38, and D33, and together with the MM-GBSA analysis revealed the hotspots leverage synergistic electrostatic and hydrophobic binding interactions in stabilizing the KRAS-RAF1 complexes. Network-based analysis of allosteric communication identifies critical KRAS residues (e.g., L6, E37, D57, R97) that mediate long-range interactions between the KRAS core and the RAF1 binding interface. The central {beta}-sheet of KRAS emerges as a hub for transmitting conformational changes, linking distant functional sites and facilitating allosteric regulation. Strikingly, the predicted allosteric hotspots align with experimentally identified allosteric binding hotspots that define the energy landscape of KRAS allostery. This study highlights the power of integrating computational modeling with experimental data to unravel the complex dynamics of KRAS and its mutants. The identification of binding hotspots and allosteric communication routes offers new opportunities for developing targeted therapies to disrupt KRAS-RAF1 interactions and inhibit oncogenic signaling. Our results underscore the potential of computational approaches to guide the design of allosteric inhibitors and mutant-specific therapies for KRAS-driven cancers.

biophysics↗

bioRxiv2024

Structure-based Design of Chimeric Influenza Hemagglutinins to Elicit Cross-group Immunity

Antigenic variability among influenza virus strains poses a significant challenge to developing broadly protective, long-lasting vaccines. Current annual vaccines target specific strains, requiring accurate prediction for effective neutralization. Despite sequence diversity across phylogenetic groups, the hemagglutinin (HA) head domains structure remains highly conserved. Utilizing this conservation, we designed cross-group chimeric HAs that combine antigenic surfaces from distant strains. By structure-guided transplantation of receptor-binding site (RBS) residues, we displayed an H3 RBS on an H1 HA scaffold. These chimeric immunogens elicit cross-group polyclonal responses capable of neutralizing both base and distal strains. Additionally, the chimeras integrate heterotrimeric immunogens, enhancing modular vaccine design. This approach enables the inclusion of diverse strain segments to generate broad polyclonal responses. In the future, such modular immunogens may serve as tools for evaluating immunodominance and refining immunization strategies, offering potential to bridge and enhance immune responses in individuals with pre-existing immunity. This strategy holds promise for advancing universal influenza vaccine development. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=105 SRC="FIGDIR/small/628867v2_ufig1.gif" ALT="Figure 1"> View larger version (25K): org.highwire.dtl.DTLVardef@18dec5borg.highwire.dtl.DTLVardef@98587forg.highwire.dtl.DTLVardef@1d9fb2dorg.highwire.dtl.DTLVardef@1f97884_HPS_FORMAT_FIGEXP M_FIG Graphical abstract: Overview of cross-group RBS transplantation approachPhylogenetically diverse HA strains can be incorporated into chimeric immunogens by RBS transplantation. The chimera are evaluated for cross-reactivity to subtype-specific antibodies and the ability to elicit neutralizing antibodies to multiple strains. C_FIG

biochemistry↗

bioRxiv2024

Diverse priming outcomes under conditions of very rare precursor B cells

Rare B cells can have special pathogen-recognition features giving them the potential to make outsized contributions to protective immunity. However, rare naive B cells infrequently participate in immune responses. We investigated how germline-targeting vaccine antigen delivery and adjuvant selection affect priming of exceptionally rare BG18-like HIV broadly neutralizing antibody-precursor B cells (~1 in 50 million) in non-human primates. Only escalating dose (ED) priming immunization using the saponin adjuvant SMNP elicited detectable BG18-like cells in germinal centers (GCs). All groups had strong GC responses, but only ED+SMNP and bolus+SMNP induced BG18-like memory B cells in >50% of animals. One group had vaccine-specific GC responses equivalent to ED+SMNP, but BG18-like memory B cells were rarely detected. Following homologous boosting, BG18-like memory B cells were more frequent in a bolus priming group, but had lower somatic hypermutation and affinities. This outcome was inversely associated with post-prime antibody titers, suggesting antibody feedback can significantly influence rare precursor B cell responses.

immunology↗

bioRxiv2024

A Multi-Property Optimizing Generative Adversarial Network for de novo Antimicrobial Peptide Design

Antimicrobial peptides (AMPs) play a crucial role in developing novel antiinfective drugs due to their broad-spectrum antimicrobial activity and lower likelihood of causing bacterial resistance. However, laboratory synthesis of AMPs is tedious and time-consuming. Existing computational methods have limited capability in optimizing multiple desired properties simultaneously. Here, we propose a Multi-Property Optimizing Generative Adversarial Network (MPOGAN), a feedback-loop framework that iteratively learns from data with multiple desired properties. This approach enables de novo design of AMPs with potent antimicrobial activity, reduced cytotoxicity, and diversity. Through extensive computational tests, MPOGAN exhibits superior performance in optimizing multiple desired properties of generated AMPs. Ten of the most promising candidates are chemically synthesized, with nine showing potent antimicrobial activity against three bacterial strains and low cytotoxicity against eukaryotic cells. MPOGAN offers a powerful computational approach for effective multi-property optimization of AMPs, thereby advancing the field of AI-aided drug discovery.

bioinformatics↗

bioRxiv2024

Mild focal cooling selectively impacts computations in dendritic trees

Focal cooling is a powerful technique to temporally scale neural dynamics. However, the underlying cellular mechanisms causing this scaling remain unresolved. Here, using targeted focal cooling (with a spatial resolution of 100 micrometers), dual somato-dendritic patch clamp recordings, two-photon calcium imaging, transmitter uncaging, and modeling we reveal that a 5{degrees}C drop can enhance synaptic transmission, plasticity, and input-output transformations in the distal apical tuft, but not in the basal dendrites of intrinsically bursting L5 pyramidal neurons. This enhancement depends on N-methyl-D-aspartate (NMDA) and Kv4.2, suggesting electrical structure modulation. Paradoxically, and despite the increase in tuft excitability, we observe a reduced rate of recovery from inactivation for apical Na+ channels, thereby regulating back-propagating action potential invasion, coincidence detection, and overall burst probability, resulting in an "apparent" slowing of somatic spike output. Our findings reveal a differential temperature sensitivity along the basal-tuft axis of L5 neurons analog modulates cortical output.

neuroscience↗

bioRxiv2024

Ultrasensitive in vivo infrared spectroscopic imaging via oblique photothermal microscopy

In vivo IR spectroscopy faces challenges due to poor sensitivity in reflection mode and low resolution at micrometer scale. To break this barrier, we report an oblique photothermal microscope (OPTM) to enable ultrasensitive IR spectroscopic imaging of live subjects at sub-micron resolution. Classic photothermal measurement captures only a small fraction of probe photons through a pinhole to extract the photothermal signal. Instead, OPTM uses a differential split detector placed on the sample surface to collect 500-fold more photons and suppress the laser noise by 12 fold via balanced detection. Leveraging its improved sensitivity, OPTM enables low-dose IR imaging of skin without photodamage. Depth-resolved in vivo OPTM imaging of metabolic markers beneath mouse and human skin is shown. Furthermore, we demonstrate in vivo OPTM tracking of topical drug contents within mouse and human skin. Collectively, OPTM presents a highly sensitive imaging platform for in vivo and in situ molecular analysis.

bioengineering↗

bioRxiv2024

Interference with Systemic Negative Feedback Regulation as a Potential Mechanism for Nonmonotonic Dose-Responses of Endocrine-Disrupting Chemicals

BackgroundEndocrine-disrupting chemicals (EDCs) often exhibit nonmonotonic doseresponse (NMDR) relationships, posing significant challenges to health risk assessment and regulations. Several molecular mechanisms operating locally in cells have been proposed, including opposing actions via different receptors, mixed-ligand heterodimer formation, and receptor downregulation. Systemic negative feedback regulation of hormone homeostasis, which is a common feature of many endocrine systems, has also been invoked as a mechanism; however, whether and how exactly such global feedback structure may underpin NMDRs is poorly understood. ObjectivesWe hypothesize that an EDC may compete with the endogenous hormone for receptors (i) at the central site to interfere with the feedback regulation thus altering the physiological hormone level, and (ii) at the peripheral site to disrupt the hormone action; this dual-action may oppose each other, producing nonmonotonic endocrine effects. The objective here is to explore - through computational modeling - how NMDRs may arise through this potential mechanism and the relevant biological variabilities that enable susceptibility to nonmonotonic effects. MethodsWe constructed a dynamical model of a generic hypothalamic-pituitary-endocrine (HPE) axis with negative feedback regulation between a pituitary hormone and a terminal effector hormone (EH). The effects of model parameters, including receptor binding affinities and efficacies, on NMDR were examined for EDC agonists and antagonists. Monte Carlo human population simulations were then conducted to systemically explore biological parameter conditions that engender NMDR. ResultsWhen an EDC interferes sufficiently with the central feedback action of EH, the net endocrine effect at the peripheral target site can be opposite to what is expected of an agonist or antagonist at low concentrations. J/U or Bell-shaped NMDRs arise when the EDC has differential binding affinities and/or efficacies, relative to EH, for the peripheral and central receptors. Quantitative relationships between these biological variabilities and associated distributions were discovered, which can distinguish J/U and Bell-shaped NMDRs from monotonic responses. ConclusionsThe ubiquitous negative feedback regulation in endocrine systems can act as a universal mechanism for counterintuitive and nonmonotonic effects of EDCs. Depending on key receptor kinetic and signaling properties of EDCs and endogenous hormones, some individuals may be more susceptible to these complex endocrine effects.

systems biology↗

bioRxiv2024

Ultrack: pushing the limits of cell tracking across biological scales

Tracking live cells across 2D, 3D, and multi-channel time-lapse recordings is crucial for understanding tissue-scale biological processes. Despite advancements in imaging technology, achieving accurate cell tracking remains challenging, particularly in complex and crowded tissues where cell segmentation is often ambiguous. We present Ultrack, a versatile and scalable cell-tracking method that tackles this challenge by considering candidate segmentations derived from multiple algorithms and parameter sets. Ultrack employs temporal consistency to select optimal segments, ensuring robust performance even under segmentation uncertainty. We validate our method on diverse datasets, including terabyte-scale developmental time-lapses of zebrafish, fruit fly, and nematode embryos, as well as multi-color and label-free cellular imaging. We show that Ultrack achieves state-of-the-art performance on the Cell Tracking Challenge and demonstrates superior accuracy in tracking densely packed embryonic cells over extended periods. Moreover, we propose an approach to tracking validation via dual-channel sparse labeling that enables high-fidelity ground truth generation, pushing the boundaries of long-term cell tracking assessment. Our method is freely available as a Python package with Fiji and napari plugins and can be deployed in a high-performance computing environment, facilitating widespread adoption by the research community.

developmental biology↗

bioRxiv2024

Large-scale genome analyses provide insights into Hymenoptera evolution

The order Hymenoptera includes a large number of species with diverse lifestyles and is known for its significant contributions to natural ecosystems. To better understand the evolution of this diverse order, we performed large-scale comparative genomics on 131 species from 13 superfamilies, covering most representative groups. We used these genomes to reveal an overall pattern of genomic change in terms of gene content and evolutionary rate throughout hymenopteran history. We identified genes that possibly contributed to the evolution of several key innovations, such as parasitoidism, wasp-waist, sting, and secondary phytophagy. We also discovered the distinct genomic trajectories between the clade containing major parasitoid wasps (Parasitoida) and stinging species (Aculeata) since their divergence, which are involved in many aspects of genomic change, such as rapidly evolving gene families, gene gain and loss, and metabolic pathway evolution. In addition, we explored the genomic features accompanying the three independent evolution of secondary phytophagy. Our work provides insights for understanding genome evolution and the genomic basis of diversification in Hymenoptera.

genomics↗

bioRxiv2024

Prediction of Conformational Ensembles and Structural Effects of State-Switching Allosteric Mutants in the Protein Kinases Using Comparative Analysis of AlphaFold2 Adaptations with Sequence Masking and Shallow Subsampling

Despite the success of AlphaFold2 approaches in predicting single protein structures, these methods showed intrinsic limitations in predicting multiple functional conformations of allosteric proteins and have been challenged to accurately capture of the effects of single point mutations that induced significant structural changes. We systematically examined several implementations of AlphaFold2 methods to predict conformational ensembles for state-switching mutants of the ABL kinase. The results revealed that a combination of randomized alanine sequence masking with shallow multiple sequence alignment subsampling can significantly expand the conformational diversity of the predicted structural ensembles and capture shifts in populations of the active and inactive ABL states. Consistent with the NMR experiments, the predicted conformational ensembles for M309L/L320I and M309L/H415P ABL mutants that perturb the regulatory spine networks featured the increased population of the fully closed inactive state. On the other hand, the predicted conformational ensembles for the G269E/M309L/T334I and M309L/L320I/T334I triple ABL mutants that share activating T334I gate-keeper substitution are dominated by the active ABL form. The proposed adaptation of AlphaFold can reproduce the experimentally observed mutation-induced redistributions in the relative populations of the active and inactive ABL states and capture the effects of regulatory mutations on allosteric structural rearrangements of the kinase domain. The ensemble-based network analysis complemented AlphaFold predictions by revealing allosteric mediating centers that often directly correspond to state-switching mutational sites or reside in their immediate local structural proximity, which may explain the global effect of regulatory mutations on structural changes between the ABL states. This study suggested that attention-based learning of long-range dependencies between sequence positions in homologous folds and deciphering patterns of allosteric interactions may further augment the predictive abilities of AlphaFold methods for modeling of alternative protein sates, conformational ensembles and mutation-induced structural transformations.

biophysics↗

bioRxiv2024

Premature thymic involution in young Foxn1lacz mutant mice causes peripheral T cell phenotypes similar to aging-induced immunosenescence

The thymus is a primary lymphoid organ generating self-restricted and self-tolerant naive T-cells. Early in life the thymus starts to involute, resulting in decreased naive T-cell output which may be more self-reactive, leading to an increased prevalence of autoimmunity. A decrease in the transcription factor FOXN1 is an early event in thymic involution. Using the Foxn1lacz model, we studied how premature thymic involution affects the thymic microenvironment, thymocytes, and peripheral T cell immunity. We found that early thymic involution led to aged-like thymic epithelial cells that resulted in aged-like thymocyte phenotypes, with a significant decrease in CD4+ single-positive T-cells. We also observed severe lymphopenia in Foxn1lacz mice caused by the premature decrease in T-cell production, resulting in a peripheral T-cell phenotype similar to de novo aged peripheral T-cells. Moreover, following T cell receptor stimulation, Foxn1lacz peripheral T cells had reduced IL-2 secretion and strong initial IFN-g responses, resembling aged wild-type peripheral T-cell responses. Lastly, influenza response in Foxn1lacz had a reduction in some aspects of T cell responses to influenza infection, similar to thymectomized mice. Our study shows an independent and direct impact of premature thymic involution on both thymopoiesis and peripheral immune niches likely contributing to immunosenescence and inflammaging as observed in the elderly population.

immunology↗

bioRxiv2024

SLC17 transporters mediate renal excretion of Lac-Phe in mice and humans

N-lactoyl-phenylalanine (Lac-Phe) is a lactate-derived metabolite that suppresses food intake and body weight. Little is known about the mechanisms that mediate Lac-Phe transport across cell membranes. Here we identify SLC17A1 and SLC17A3, two kidney-restricted plasma membrane-localized solute carriers, as physiologic urine Lac-Phe transporters. In cell culture, SLC17A1/3 exhibit high Lac-Phe efflux activity. In humans, levels of Lac-Phe in urine exhibit a strong genetic association with the SLC17A1-4 locus. Urine Lac-Phe levels are also increased following a Wingate sprint test. In mice, genetic ablation of either SLC17A1 or SLC17A3 reduces urine Lac-Phe levels. Despite these differences, both knockout strains have normal blood Lac-Phe and body weights, demonstrating that urine and plasma Lac-Phe pools are functionally and biochemically de-coupled. Together, these data establish SLC17 family members as the physiologic urine transporters for Lac-Phe and uncover a biochemical pathway for the renal excretion of this signaling metabolite.

biochemistry↗

bioRxiv2024

Predicting Functional Conformational Ensembles and Binding Mechanisms of Convergent Evolution for SARS-CoV-2 Spike Omicron Variants Using AlphaFold2 Sequence Scanning Adaptations and Molecular Dynamics Simulations

In this study, we combined AlphaFold-based approaches for atomistic modeling of multiple protein states and microsecond molecular simulations to accurately characterize conformational ensembles and binding mechanisms of convergent evolution for the SARS-CoV-2 Spike Omicron variants BA.1, BA.2, BA.2.75, BA.3, BA.4/BA.5 and BQ.1.1. We employed and validated several different adaptations of the AlphaFold methodology for modeling of conformational ensembles including the introduced randomized full sequence scanning for manipulation of sequence variations to systematically explore conformational dynamics of Omicron Spike protein complexes with the ACE2 receptor. Microsecond atomistic molecular dynamic simulations provide a detailed characterization of the conformational landscapes and thermodynamic stability of the Omicron variant complexes. By integrating the predictions of conformational ensembles from different AlphaFold adaptations and applying statistical confidence metrics we can expand characterization of the conformational ensembles and identify functional protein conformations that determine the equilibrium dynamics for the Omicron Spike complexes with the ACE2. Conformational ensembles of the Omicron RBD-ACE2 complexes obtained using AlphaFold-based approaches for modeling protein states and molecular dynamics simulations are employed for accurate comparative prediction of the binding energetics revealing an excellent agreement with the experimental data. In particular, the results demonstrated that AlphaFold-generated extended conformational ensembles can produce accurate binding energies for the Omicron RBD-ACE2 complexes. The results of this study suggested complementarities and potential synergies between AlphaFold predictions of protein conformational ensembles and molecular dynamics simulations showing that integrating information from both methods can potentially yield a more adequate characterization of the conformational landscapes for the Omicron RBD-ACE2 complexes. This study provides insights in the interplay between conformational dynamics and binding, showing that evolution of Omicron variants through acquisition of convergent mutational sites may leverage conformational adaptability and dynamic couplings between key binding energy hotspots to optimize ACE2 binding affinity and enable immune evasion.

biophysics↗

bioRxiv2024

A PTER-dependent pathway of taurine metabolism linked to energy balance

Taurine is a conditionally essential micronutrient and one of the most abundant amino acids in humans1-3. In endogenous taurine metabolism, dedicated enzymes are involved in biosynthesis of taurine from cysteine as well as the downstream derivatization of taurine into secondary taurine metabolites4,5. One such taurine metabolite is N-acetyltaurine6. Levels of N-acetyltaurine are dynamically regulated by diverse physiologic perturbations that alter taurine and/or acetate flux, including endurance exercise7, nutritional taurine supplementation8, and alcohol consumption6,9. While taurine N-acetyltransferase activity has been previously detected in mammalian cells6,7, the molecular identity of this enzyme, and the physiologic relevance of N-acetyltaurine, have remained unknown. Here we show that the orphan body mass index-associated enzyme PTER (phosphotriesterase-related)10 is the principal mammalian taurine N-acetyltransferase/hydrolase. In vitro, recombinant PTER catalyzes bidirectional taurine N-acetylation with free acetate as well as the reverse N-acetyltaurine hydrolysis reaction. Genetic ablation of PTER in mice results in complete loss of tissue taurine N-acetyltransferase/hydrolysis activities and systemic elevation of N-acetyltaurine levels. Upon stimuli that increase taurine levels, PTER-KO mice exhibit lower body weight, reduced adiposity, and improved glucose homeostasis. These phenotypes are recapitulated by administration of N-acetyltaurine to wild-type mice. Lastly, the anorexigenic and anti-obesity effects of N-acetyltaurine require functional GFRAL receptors. Together, these data uncover enzymatic control of a previously enigmatic pathway of secondary taurine metabolism linked to energy balance.

biochemistry↗

bioRxiv2024

Ovarian disrupting effects and mechanisms of long- and short-chain per- and polyfluoroalkyl substances in mice

BackgroundThe extensive use of per- and polyfluoroalkyl substances (PFAS) has led to environmental contamination and bioaccumulation. Previous research linked PFAS exposure to female reproductive disorders, but the mechanism remains elusive. Further, most studies focused on legacy long-chain PFOA and PFOS, yet the reproductive impacts of other long-chain PFAS and short-chain alternatives are rarely explored. ObjectivesWe investigated the effects and mechanisms of long- and short-chain PFAS on the ovary and associated ovarian functions. MethodsA 3D in vitro ovarian follicle culture system and an in vivo mouse model, together with approaches of reverse transcription-quantitative polymerase chain reaction, enzyme-linked immunosorbent assay, RNA-sequencing, pharmacological treatment, in situ zymography, histology, in situ hybridization, analytical chemistry, and benchmark dose modeling (BMD), were used to test environmentally relevant exposure levels of six long- and short-chain PFAS on follicle maturation, hormone secretion, and ovulation. ResultsIn vitro exposure revealed that long-but not short-chain PFAS interfered with gonadotropin-dependent follicle maturation, ovulation, and hormone secretion. Mechanistically, long-chain perfluorononanoic acid (PFNA) acted as a peroxisome proliferator-activated receptor gamma (PPAR{gamma}) agonist in granulosa cells to disrupt follicle-stimulating hormone (FSH)-dependent follicle maturation, luteinizing hormone (LH)-stimulated ovulation, and associated gene regulatory pathways. In vivo mouse exposure confirmed the ovarian accumulation of PFNA and the mechanism of PPAR{gamma}-mediated ovarian toxicities of PFNA observed in vitro. The BMD analysis of in vitro and in vivo results suggested human relevant exposure levels of long-chain PFAS in our study pose an extra risk of ovarian defects, with follicular rupture as the most sensitive endpoint. DiscussionUsing in vitro follicle culture and in vivo mouse models, we discovered that long-chain PFAS interfere with gonadotropin-dependent follicle maturation, hormone secretion, and ovulation, posing a non-negligible risk to womens reproductive health including anovulation, irregular menstrual cycles, and sub- or infertility.

pharmacology and toxicology↗

bioRxiv2024

Dose-response functional and transcriptomic effects of follicle-stimulating hormone on ex vivo mouse folliculogenesis

The gonadotropin-dependent phase of ovarian folliculogenesis primarily requires follicle-stimulating hormone (FSH) to support one or multiple antral follicles, dependent on the species, to mature fully, enabling ovarian steroidogenesis, oogenesis, and ovulation to sustain female reproductive cycles and fertility. FSH binds to its membrane receptor in granulosa cells to activate various signal transduction pathways and gene regulatory networks. Poor female reproductive outcomes can result from both FSH insufficiency owing to genetic or non-genetic factors and FSH excess as encountered with ovarian stimulation in assisted reproductive technology (ART), but the underlying molecular mechanisms remain elusive. Herein, we conducted single-follicle and single-oocyte RNA sequencing analysis along with other approaches in an ex vivo mouse folliculogenesis and oogenesis system to investigate the effects of different concentrations of FSH on key follicular events. Our study revealed that a minimum FSH threshold is required for follicle maturation into the high estradiol-secreting preovulatory stage, and the threshold is moderately variable among individual follicles. FSH at subthreshold, threshold, and suprathreshold levels induced distinct expression patterns of follicle maturation-related genes and the follicular transcriptomics. The RNA-seq analysis identified novel genes and signaling pathways that may critically regulate follicle maturation. Suprathreshold FSH resulted in multiple ovarian disorders including premature luteinization, high production of androgen and proinflammatory factors, and reduced expression of energy metabolism-related genes in oocytes. Together, this study improves our understanding of gonadotropin-dependent folliculogenesis and provides crucial insights into how high doses of FSH used in ART may impact follicular health, oocyte quality, pregnancy outcome, and systemic health.

molecular biology↗

bioRxiv2024

Interpretable Atomistic Prediction and Functional Analysis of Conformational Ensembles and Allosteric States in Protein Kinases Using AlphaFold2 Adaptation with Randomized Sequence Scanning and Local Frustration Profiling

The groundbreaking achievements of AlphaFold2 (AF2) approaches in protein structure modeling marked a transformative era in structural biology. Despite the success of AF2 tools in predicting single protein structures, these methods showed intrinsic limitations in predicting multiple functional conformations of allosteric proteins and fold-switching systems. The recent NMR-based structural determination of the unbound ABL kinase in the active state and two inactive low-populated functional conformations that are unique for ABL kinase presents an ideal challenge for AF2 approaches. In the current study we employ several implementations of AF2 methods to predict protein conformational ensembles and allosteric states of the ABL kinase including (a) multiple sequence alignments (MSA) subsampling approach; (b) SPEACH_AF approach in which alanine scanning is performed on generated MSAs; and (c) introduced in this study randomized full sequence mutational scanning for manipulation of sequence variations combined with the MSA subsampling. We show that the proposed AF2 adaptation combined with local frustration mapping of conformational states enable accurate prediction of the ABL active and intermediate structures and conformational ensembles, also offering a robust approach for interpretable characterization of the AF2 predictions and limitations in detecting hidden allosteric states. We found that the large high frustration residue clusters are uniquely characteristic of the low-populated, fully inactive ABL form and can define energetically frustrated cracking sites of conformational transitions, presenting difficult targets for AF2 methods. This study uncovered previously unappreciated, fundamental connections between distinct patterns of local frustration in functional kinase states and AF2 successes/limitations in detecting low-populated frustrated conformations, providing a better understanding of benefits and limitations of current AF2-based adaptations in modeling of conformational ensembles.

biophysics↗

bioRxiv2024

Genomic insights into photosymbiotic evolution in Tridacna squamosa

Photosymbiosis is fundamental driving force for ecological success of benthic coral reef ecosystems, and contributing to their biodiversity and resilience. As a benchmark organism indicative of reef health, the fluted giant clam (Tridacna squamosa) forms an exemplary photosymbiotic relationship with the symbiont Symbiodiniaceae dinoflagellates, whose initiation and maturation require finely coordinated interactions. However, much of the origin and dynamics of this reciprocal interplay remains unclarified. Here, we report the first complete whole genome of T. squamosa, in conjunction with integrated multi-omics data, to illuminate the key evolutionary innovations and molecular events supporting the establishment and maintenance of photosymbiotic lifestyle in the giant clam. Programmed regulation of symbiont recognition, host immune system and GPCRs signaling activation co-contributed to dinoflagellates acquisition in T. squamosa larvae. Adaptive metabolic remodeling in the host siphonal mantle, a photosymbiotic niche, is critical to maintain the robustness of phtosymbiosis. T. squamosa has expanded light sensing gene family and evolved sophisticated signaling pathways to protect against UV photo-damage. Evidence also supports significant contribution of positive selection to host DNA-repair. Overall, our study here offers fresh mechanistic insights into the parallel evolution and molecular machinery of photosymbiosis in the giant clam-dinoflagellates duet, with implications for devising solutions to sustainable conservation.

evolutionary biology↗