Biology subjects

Wang, F.

Publications and source records attributed to Wang, F..

At least 217 records · Page 12Linked to original sources

bioRxiv2024

Self-Assembled Chambered Cardiac Organoids for Modeling Cardiac Chamber Formation and Cardiotoxicity Assessment

The human heart, crucial for health and longevity, is a primary focus of medical research. Despite advancements in cardiac organoid technology, replicating early cardiac chamber formation stages remains challenging. Here, we develop chambered cardiac organoids (CCOs) by orchestrating cardiac signaling pathways, including the synergistic modulation of FGF and Wnt signaling via the HAND1 transcription factor. These CCOs exhibit stable chambers with self-organized outer myocardial and inner endocardial layers, and express developmental markers such as NKX2.5, TNNT2, and NAPPA, demonstrating physiological functionality including spontaneous contractions and calcium transients. Immunofluorescence and single-cell RNA sequencing confirmed the presence and stability of cardiomyocytes within CCOs. Further validation showed CCOs involvement in critical processes like endothelial-mesenchymal transition (EndoMT) and valvulogenesis. Ultrastructural and electrophysiological analysis revealed organized myofibrils and atrial-like action potentials. Importantly, CCOs proved effective in assessing cardiotoxicity through observable morphological changes, demonstrating specific cellular responses to established cardiotoxic compounds, thereby highlighting their potential for drug testing and safety evaluations. Our findings offer insights into cardiac chamber formation mechanisms and establish a robust platform for drug testing, disease modeling, and personalized medicine, representing a significant advancement in functional cardiac organoid development for both basic research and translational applications.

cell biology↗

bioRxiv2024

Noncanonical roles of ATG5 and membrane atg8ylation in retromer assembly and function

ATG5 is one of the core autophagy proteins with additional functions such as noncanonical membrane atg8ylation, which among a growing number of biological outputs includes control of tuberculosis in animal models. Here we show that ATG5 associates with retromers core components VPS26, VPS29 and VPS35 and modulates retromer function. Knockout of ATG5 blocked trafficking of a key glucose transporter sorted by the retromer, GLUT1, to the plasma membrane. Knockouts of other genes essential for membrane atg8ylation, of which ATG5 is a component, affected GLUT1 sorting, indicating that membrane atg8ylation as a process affects retromer function and endosomal sorting. The contribution of membrane atg8ylation to retromer function in GLUT1 sorting was independent of canonical autophagy. These findings expand the scope of membrane atg8ylation to specific sorting processes in the cell dependent on the retromer and its known interactors.

cell biology↗

bioRxiv2024

TgATG9 is required for autophagosome biogenesis and maintenance of chronic infection in Toxoplasma gondii

Toxoplasma gondii is a ubiquitous protozoan parasite that can reside long-term within hosts as intracellular tissue cysts comprised of chronic stage bradyzoites. To perturb chronic infection requires a better understanding of the cellular processes that mediate parasite persistence. Macroautophagy/autophagy is a catabolic and homeostatic pathway that is required for T. gondii chronic infection, although the molecular details of this process remain poorly understood. A key step in autophagy is the initial formation of the phagophore that sequesters cytoplasmic components and matures into a double-membraned autophagosome for delivery of the cargo to a cells digestive organelle for degradative recycling. While T. gondii appears to have a reduced repertoire of autophagy proteins, it possesses a putative phospholipid scramblase, TgATG9. Through structural modeling and complementation assays, we show herein that TgATG9 can partially rescue bulk autophagy in atg9{Delta} yeast. We demonstrated the importance of TgATG9 for proper autophagosome dynamics at the subcellular level using three-dimensional live cell lattice light sheet microscopy. Conditional knockdown of TgATG9 in T. gondii after bradyzoite differentiation resulted in markedly reduced parasite viability. Together, our findings provide insights into the molecular dynamics of autophagosome biogenesis within an early-branching eukaryote and pinpoint the indispensable role of autophagy in maintaining T. gondii chronic infection.

microbiology↗

bioRxiv2024

Activation of two noncanonical R proteins by an insect effector confers plant immunity to aphid infestation

Molecular characterization of resistance genes is crucial for efficiently understanding and fortifying plant immunity against insect herbivores. Here we report that RMES1A and RMES1B proteins confer resistance to the sorghum aphid Melanaphis sorghi when activated by an insect effector MsEF1. Map-based cloning plus genetic analysis of knockout mutants confirm that RMES1A and RMES1B are both required for aphid resistance. Upon aphid attack, RMES1A and RMES1B expression is elevated in the sclerenchyma cells and vascular bundles of leaves; the two proteins interact with MsEF1 in the exocysts, thus upregulating key defense processes such as reactive oxygen species burst. Structural modeling predicts that RMES1A and RMES1B each carry an ATP binding site and two leucine-rich-repeat domains but lack coiled-coil or Toll/Interleukin-1 receptor/resistance domain, thus likely representing a new type of resistance controlling proteins in plants. Our work reveals new genes and mechanisms for further deciphering and improving plant immunity to insect pests.

genetics↗

bioRxiv2024

Functional analysis of ESRP1/2 gene variants and CTNND1 isoforms in orofacial cleft pathogenesis

Orofacial cleft (OFC) is a common human congenital anomaly. Epithelial-specific RNA splicing regulators ESRP1 and ESRP2 regulate craniofacial morphogenesis and their disruption result in OFC in zebrafish, mouse and humans. Using esrp1/2 mutant zebrafish and murine Py2T cell line models, we functionally tested the pathogenicity of human ESRP1/2 gene variants. We found that many variants predicted by in silico methods to be pathogenic were functionally benign. Esrp1 also regulates the alternative splicing of Ctnnd1 and these genes are co-expressed in the embryonic and oral epithelium. In fact, over-expression of ctnnd1 is sufficient to rescue morphogenesis of epithelial-derived structures in esrp1/2 zebrafish mutants. Additionally, we identified 13 CTNND1 variants from genome sequencing of OFC cohorts, confirming CTNND1 as a key gene in human OFC. This work highlights the importance of functional assessment of human gene variants and demonstrates the critical requirement of Esrp-Ctnnd1 acting in the embryonic epithelium to regulate palatogenesis.

genetics↗

bioRxiv2024

CD-GPT: A Biological Foundation Model Bridging the Gap between Molecular Sequences Through Central Dogma

The central dogma serves as a fundamental framework for understanding the flow and expression of genetic information within living organisms, facilitating the connection of diverse biological sequences across molecule types. In this study, we present CD-GPT (Central Dogma Generative Pretrained Transformer), a generative biological foundation model with 1 billion parameters, aiming to capture the sequence relationships between DNA, RNA, and proteins. We model sequences in a unified representational space and employ a shared, multi-molecule vocabulary to narrow their distances in the embedding space effectively. Through extensive pretraining on nucleotide and amino acid sequence data, CD-GPT exhibits exceptional performance in a wide range of predictive and generative downstream tasks, including mono-molecular and multi-molecular analyses. Notably, CD-GPT excels in tasks such as genomic element detection, protein property prediction, RNA-protein interaction identification and also generative tasks like protein generation and reverse translation. The versatility of CD-GPT opens up promising avenues for advanced multi-omics analysis.

bioinformatics↗

bioRxiv2024

In a nutshell: pistachio genome and kernel development

Pistachio is a sustainable nut crop with exceptional climate resilience and nutritional value. To advance pistachios as a future food source and a model system for hard-shelled fruits, we generated a chromosome-scale reference genome of the most widely grown pistachio cultivar (Pistacia vera Kerman) and a spatiotemporal developmental study of the hull, shell, and kernel. Our study defined four distinct stages of pistachio growth and maturation by integrating tissue-level physiological and molecular data from thousands of nuts across twenty-four time points over three growing seasons. Transcriptional and metabolic changes in the kernel elucidate molecular pathways governing nutritional quality, such as the accumulation of unsaturated fatty acids, which are vital for shelf-life and dietary value. This work yields new knowledge and resources that will inform other woody crops and facilitate further improvement of pistachio as a globally significant, sustainable, and nutritious crop.

plant biology↗

bioRxiv2024

Identification of Multi-landscape and Cell Interactions in the Tumor Microenvironment through High-Coverage Single-Cell Sequencing

Single-cell RNA sequencing (scRNA-seq) is a widely used method for classifying cell types and states and revealing disease mechanisms. However, most contemporary scRNA-seq platforms fail to explore the multilandscape of RNA. Here, we designed a microfluidic chip that combines oligo-dT primers and Random Bridging Co-labeling (RBCL) RNA sequencing to develop an innovative Chigene scRNA-seq technology that can identify gene expression, mutations, and RNA splicing landscapes at the single-cell level. The Chigene scRNA-seq platform demonstrated exceptional performance, with minimal doublet rates of 0.94% (Chigene V1) and 1.93% (Chigene V2). Both versions exhibit high sensitivity, with Chigene V2 achieving nearly 100% RNA coverage and detecting over 1800 genes per cell on average. Targeted capture of single-cell gene mutations enhances mutation detection sensitivity. Moreover, this Chigene V2 platform has been validated in clinical samples for its ability to detect mutations, gene fusions and alternative splicing. The reliability of the platform was further corroborated via known functional gene mutation (CDKN1A) and fusion (FGFR3-TACC). To validate this methods potential for discovering novel gene mutations in clinical samples, our investigation revealed an intriguing cell subpopulation carrying an ARHGAP5 mutation in urothelial carcinoma. These cells exhibited high-frequency mRNA splicing and exhibited specific crosstalk with T cells, distinguishing them from the subpopulation with the ARHGAP5 wild-type phenotype. Overall, this method provides a robust scRNA-seq platform suitable for comprehensive analyses of clinical specimens at different genetic information levels, thereby offering significant potential in the discovery of novel genes and interactions at the single-cell level.

genomics↗

bioRxiv2024

RCOR1 promotes myoblast differentiation and muscle regeneration

RCOR proteins belong to a family of highly conserved transcription corepressors (RCOR1, RCOR2 and RCOR3) that regulate the activity of associated histone demethylase 1 (LSD1) and histone deacetylase 1/2 (HDAC 1/2) in chromatin-modifying complexes. Despite the described function of LSD1 in skeletal muscle differentiation and regeneration, the role of RCOR family in myogenesis remains unknown. We found that RCOR1 is highly expressed in proliferating myoblasts and activated satellite cells, but not in mature myofibers during postnatal growth and regeneration of skeletal muscle. Knockdown of RCOR1 impaired myoblast differentiation and fusion by inhibiting the expression of the key myogenic regulatory factor myogenin. Moreover, RCOR1 depletion impaired myoblast proliferation through increasing the expression of cell cycle inhibitor p21. Consistently, in a mouse model of skeletal muscle injury, depletion of RCOR1 supressed satellite cell activation and differentiation which resulted in impaired muscle regeneration. RCOR1 was found physically associated with LSD1 and myogenic regulatory factor MyoD and contributed to LSD1 stability in myoblasts. As for other RCOR family members, RCOR2 had no effect on myoblast differentiation while the loss of RCOR3 increased myoblast proliferation leading to supressed expression of myogenic markers MyoD and myogenin and impaired myoblast differentiation. However, germline deletion of RCOR3 (RCOR3-/-) did not affect muscle phenotype, suggesting a possible functional redundancy among RCOR family members during muscle development. Together, our findings indicate that RCOR1 acts in concert with LSD1 as a novel positive regulator of myogenesis and skeletal muscle regeneration.

physiology↗

bioRxiv2024

Production and functional verification of 8-gene (GGTA1, CMAH, β4GalNT2, hCD46, hCD55, hCD59, hTBM, hCD39)-edited donor pigs for xenotransplantation

Gene-edited pig-to-human xenotransplantation continues to make breakthroughs and is expected to enter clinic to solve the global shortage of donor organs. However, which gene combination is suitable for which organ transplantation remains unclear. In this study, we utilized CRISPR/Cas9 gene editing technology, PiggyBac transposon system and somatic cell cloning to construct GTKO/CMAHKO/{beta}4GalNT2KO/hCD46/hCD55/hCD59/hCD39/hTBM 8 gene-edited cloned (GEC) donor pigs, and performed pig to non-human primate (NHP) transplantation to evaluate the effectiveness of these GEC pigs. The multiple vectors were co-transfected into fetal fibroblasts of Diannan miniature pig with O blood type, and 25 colonies were screened out, and one of them carried GGTA1, CMAH and {beta}4GalNT2 biallelic knockout and integration of hCD46, hCD55, hCD59, hTBM and hCD39 genes, which was used as a donor cell for cloning, and a 33-day-old viable fetus was obtained. The fetus was identified and confirmed for normal karyotype and the absence of three xenogeneic antigens -Gal, Neu5Gc and Sda, and expression of hCD46, hCD55, hCD59, hTBM and hCD39 genes, then the recloning was carried out and 28 cloned piglets were obtained by natural delivery. Molecular identification at DNA, mRNA and protein levels showed that 8 gene editing (GE) was successful in these GEC piglets. Moreover, antigen-antibody binding assay and complement-dependent cytotoxicity assay demonstrated that 8GE effectively reduced the immune incompatibility and kidney xenograft survived up to 15 and 17 days into two NHPs, respectively. During this period, the recipient serum antibodies IgA and IgM, complements C3 and C4, coagulation indicators PT, APTT, TT and FIB, as well as most electrolytes and liver function indicators remained relatively stable. The 24-hour urine output and serum creatinine remained normal at a period of post-transplantation. These results indicated that the 8GEC pigs effectively alleviated immune rejection and exerted life-supporting kidney function in the recipient.

genetics↗

bioRxiv2024

Compositional Shifts and Co-occurrence Patterns of Topsoil Bacteria and Micro-Eukaryotes Across a Permafrost Thaw Gradient in Alpine Meadows of the Qilian Mountains, China

Soil microorganisms play a pivotal role in the biogeochemical cycles of alpine meadow ecosystems, especially in the context of permafrost thaw. However, the mechanisms driving microbial community responses to environmental changes, such as variations in active layer thickness (ALT) of permafrost, remain poorly understood. This study utilized next-generation sequencing to explore the composition and co-occurrence patterns of soil microbial communities, focusing on bacteria and micro-eukaryotes along a permafrost thaw gradient. Results showed a decline in bacterial alpha diversity with increasing permafrost thaw, while micro-eukaryotic diversity exhibited an opposite trend. Although changes in microbial community composition were observed in permafrost and seasonally frozen soils, these shifts were not statistically significant. Bacterial communities exhibited greater differentiation between frozen and seasonally frozen soils, a pattern not mirrored in eukaryotic communities. LEfSe analysis revealed a higher number of potential biomarkers in bacterial communities compared to micro-eukaryotes. Bacterial co-occurrence networks were more complex, with more nodes, edges, and positive linkages than those of micro-eukaryotes. Key factors such as soil texture, ALT, and bulk density significantly influenced bacterial community structures, particularly affecting the relative abundances of the Acidobacteria, Proteobacteria, and Actinobacteria phyla. In contrast, fungal communities (e.g., Nucletmycea, Rhizaria, Chloroplastida, and Discosea groups) were more affected by electrical conductivity, vegetation coverage, and ALT. This study highlights the distinct responses of soil bacteria and micro-eukaryotes to permafrost thaw, offering insights into microbial community stability under global climate change. ImportanceThis study sheds light on how permafrost thaw affects microbial life in the soil, with broader implications for understanding climate change impacts. As permafrost degrades, it alters the types and numbers of microbes in the soil. These microbes play essential roles in environmental processes, such as nutrient cycling and greenhouse gas emissions. By observing shifts from bacteria-dominated to fungi-dominated communities as permafrost thaws, the study highlights potential changes in these processes. Importantly, this research suggests that the stability of microbial networks decreases with permafrost degradation, potentially disrupting the delicate balance of these ecosystems. The findings not only deepen our understanding of microbial responses to changing climates but also support the development of strategies to monitor and potentially mitigate the effects of climate change on fragile high-altitude ecosystems.

microbiology↗

bioRxiv2024

Silicone Wire Embolization-induced Acute Retinal Artery Ischemia and Reperfusion Model in Mouse: Gene Expression Provide Insight into Pathological Processes

Acute retinal ischemia and ischemia-reperfusion injury are the primary causes of retinal neural cell death and vision loss in retinal artery occlusion (RAO). The absence of an accurate mouse model for simulating the retinal ischemic process has hindered progress in developing neuroprotective agents for RAO. We developed a unilateral pterygopalatine ophthalmic artery occlusion (UPOAO) mouse model using silicone wire embolization combined with carotid artery ligation. The survival of retinal ganglion cells and visual function were evaluated to determine the duration of ischemia. Immunofluorescence staining, optical coherence tomography, and haematoxylin and eosin staining were utilized to assess changes in major neural cell classes and retinal structure degeneration at two reperfusion durations. Transcriptomics was employed to investigate alterations in the pathological process of UPOAO following ischemia and reperfusion, highlighting transcriptomic differences between UPOAO and other retinal ischemia-reperfusion models. The UPOAO model successfully replicated the acute interruption of retinal blood supply observed in RAO. 60-minutes of Ischemia led to significant loss of major retinal neural cells and visual function impairment. Notable thinning of the inner retinal layer, especially the ganglion cell layer, was evident post-UPOAO. Temporal transcriptome analysis revealed various pathophysiological processes related to immune cell migration, oxidative stress, and immune inflammation during the non-reperfusion and reperfusion periods. A pronounced increase in microglia within the retina and peripheral leukocytes accessing the retina was observed during reperfusion periods. Comparison of differentially expressed genes (DEGs) between the UPOAO and high intraocular pressure models revealed specific enrichments in lipid and steroid metabolism-related genes in the UPOAO model. The UPOAO model emerges as a novel tool for screening pathogenic genes and promoting further therapeutic research in RAO.

neuroscience↗

bioRxiv2024

Multi-layered apoplastic barrier underlying the ability of Na+ exclusion in Vigna marina

O_LISalt tolerance is important to tackle problems of soil salinization and ground water depletion. However, developing salt tolerant crops is facing difficulties due to limited potential in model plants and crop species. Thus, it is important to elucidate how coastal species, such as Vigna marina, adapt to saline environments. C_LIO_LIBy comparative transcriptome and histological analyses, this study elucidated one important aspect of how Vigna marina achieves salt exclusion and extraordinary salt tolerance. C_LIO_LIUnder salt stress, genes involved in casparian strip formation were specifically upregulated in JP247202 (V. marina). JP247202 reinforced apoplastic barrier with thick lignification in multiple layers of cells around endodermis. Also, disruption of lignification led to a dramatic increase of shoot Na+ accumulation and salt lesion in JP247202. Interestingly, despite the salt-induced apoplastic barrier, JP247202 maintained transport of essential ions including K+, Mg2+, and Ca2+. C_LIO_LIOur results revealed that lignification of multi-layered cells around endodermis was an important apoplastic barrier to the transport of Na+ to shoots in JP247202, while it did not restrict the transport of K+, Mg2+, and Ca2+. This feature, together with the ability of Na+ excretion by SOS1, enables V. marina to thrive in marine beaches. C_LI

plant biology↗

bioRxiv2024

Identification of Glucosidase II that regulates STIM1 activation dynamics

The ubiquitous presence of stromal interaction molecule (STIM), endoplasmic reticulum (ER) Ca2+ sensors, plays a crucial role for maintaining Ca2+ homeostasis and signaling in mammalian cells by linking ER Ca2+ depletion with extracellular Ca2+ influx. Although recent advancements have shed light on glycosylation in STIM proteins, further exploration of deglycosylation enzyme modification and its role remains limited by methodological constraints. In this study, leveraging the miniTurbo-driven proximal biotinylation labeling method, we were able to screen for weak and transiently binding protein molecules within specific environments like ER lumina. Herein, we unveil glucosidase II, comprising GANAB and PRKCSH, as a novel partner of the STIM1 complex. Through investigations into Glucosidase IIs (Glu II) regulation of STIM1s Ca2+ affinities both in cellulo and in situ, alongside its impact on store-operated Ca2+ entry (SOCE), we propose a novel mechanism wherein Glu II governs STIM1 activation by modulating Ca2+ binding affinity, thereby adjusting the level of activated STIM1 in response to physiological stimulation.

cell biology↗

bioRxiv2024

Generative Modelling of Oncogene-carrying Extrachromosomal Circular DNA Biogenesis and Dynamics in Cells

Extrachromosomal circular DNAs (ecDNA) are focal gene amplifications frequently associated with cancer development and often indicating a poor prognosis. To understand the early dynamics of oncogene-carrying ecDNAs, we previously developed CRISPR-C, a tool for precise ecDNA generation by deleting specific chromosomal regions. Here, we adapted CRISPR-C to recreate tumor ecDNAs. This method also allowed us to enhance ecDNA generation efficiency by directly delivering Cas9 protein and sgRNAs as a ribonucleoprotein complex. By using the modified CRISPR-C, we successfully generated ecDNAs carrying oncogenes (EGFR, CDK4, MDM2, MYC, MYCN, FGFR2, ABCB1, and DHFR) in various human cell types. Furthermore, we demonstrated that our method could generate chimeric ecDNAs composed of target sequences from distant intra or inter-chromosomal regions. Using these generative ecDNA cell models, we studied the oncogene ecDNA expression and stability. The MDM2 expression was increased after CRISPR-C, while CDK4 was decreased indicating genomic-context dependent effect. The copy number of CRISPR-C generated CDK4 was ecDNA increased in cells after a long period of treatment with the CDK4 inhibitor palbociclib. Unlike CDK4, the CRISPR-C generated ABCB1 ecDNA was unstable in cells under normal growth conditions, but is stably retained when the cells were treated with colcemid, a recognized substrate for ABCB1. We thus provide valuable tools and an attractive platform for studying ecDNA biogenesisy and in vitro drug screening on ecDNA stability.

cancer biology↗

bioRxiv2024

InstructPLM: Aligning Protein Language Models to Follow Protein Structure Instructions

Large language models are renowned for their efficacy in capturing intricate patterns, including co-evolutionary relationships, and underlying protein languages. However, current methodologies often fall short in illustrating the emergence of genomic insertions, duplications, and insertion/deletions (indels), which account for approximately 14% of human pathogenic mutations. Given that structure dictates function, mutated proteins with similar structures are more likely to persist throughout biological evolution. Motivated by this, we leverage crossmodality alignment and instruct fine-tuning techniques inspired by large language models to align a generative protein language model with protein structure instructions. Specifically, we present a method for generating variable-length and diverse proteins to explore and simulate the complex evolution of life, thereby expanding the repertoire of options for protein engineering. Our proposed protein LM-based approach, InstructPLM, demonstrates significant performance enhancements both in silico and in vitro. On native protein backbones, it achieves a perplexity of 2.68 and a sequence recovery rate of 57.51, surpassing Protein-MPNN by 39.2% and 25.1%, respectively. Furthermore, we validate the efficacy of our model by redesigning PETase and L-MDH. For PETase, all fifteen designed variable-length PETase exhibit depolymerization activity, with eleven surpassing the activity levels of the wild type. Regarding L-MDH, an enzyme lacking an experimentally determined structure, InstructPLM is able to design functional enzymes with an AF2-predicted structure. Code and model weights of InstructPLM are publicly available*.

bioinformatics↗

bioRxiv2024

Why is usefulness rarely useful?

Mate selection plays an important role in breeding programs. The usefulness criterion was proposed as a criterion for mate selection, combining information on both the mean and standard deviation of the potential offspring, particularly in clonally propagated outbred species where large family sizes are possible. Predicting mean values of offspring of a cross is generally easier than predicting the standard deviation, especially in outbred species where the linkage of alleles is often unknown and phasing is required. In this study, we developed a method for estimating phasing accuracy from unphased genotype data on possible parental lines and evaluated whether the accuracy was sufficient to predict family standard deviations of possible crosses using a set of simulations spanning a wide range of genetic architectures and genotypes from a real strawberry breeding population. We find that despite highly accurate computational phasing, predicting family standard deviations and using predicted values of the usefulness criterion per possible cross confers little benefit relative to simply selecting parents based on predicted family means. Therefore even in this species, which is clonally propagated, outbred, and produces large families, we find the usefulness criterion unlikely to be useful.

genetics↗

bioRxiv2024

Coral long-term recovery after bleaching: implications for sexual reproduction and physiology.

This study examined the long-term impacts of coral bleaching on the reproduction and physiology of Montipora capitata, a dominant reef-building coral in Hawaii. We monitored bleached and non-bleached colonies during and after a natural coral bleaching event in 2014 and analyzed reproductive traits and transcriptomic signatures eight months later. Our study shows that non-bleached and bleached colonies successfully produced gametes. Colonies that bleached had smaller oocytes, and development was slower than in colonies that did not bleach. Corals with different vulnerabilities to bleaching exhibited distinct transcriptomic responses eight months after a bleaching event. Those more prone to bleaching showed suppression of transcripts associated with sperm motility, calcification, and immunity. We found distinct transcriptomic signatures between fringing and patch reefs, suggesting local adaptation and/or acclimatization. To conserve coral reefs and better understand how they will be affected by future heat stress, we need to track which colonies survive and examine how their physiological and reproductive processes are impacted in the short- and long-term. This is critical as consecutive bleaching events become more frequent, and corals have less time to recover. Our study provides valuable molecular and reproductive data that can be used for conservation and management purposes. This information can help us identify signs of coral vulnerability and resilience to bleaching, project how future bleaching events will affect coral reproduction, determine which traits are most at risk, and assess which sites are more likely to be compromised.

physiology↗