Biology subjects

Hunter, N.

Publications and source records attributed to Hunter, N..

At least 19 recordsLinked to original sources

bioRxiv2026

Auto-sumoylation of UBC9 coordinates meiotic prophase and protects ovarian reserves

The small ubiquitin-like modifier SUMO regulates key events of meiosis, including pairing and crossing over between homologous chromosomes. Auto-sumoylation of the SUMO E2-conjugating enzyme UBC9 at lysine 14 alters its substrate selectivity in vitro, but the role of this modification in vivo is unknown. Here, we show that UBC9-K14 auto-sumoylation helps coordinate meiotic prophase and is important for maintenance of the ovarian reserve. Ubc9K14R/K14R knock-in mice show a variety of defects in meiotic prophase I, including altered assembly of DNA strand-exchange complexes, delayed and defective homolog synapsis, and unstable crossover recombination complexes. In spermatocytes, these defects are associated with reduced efficiency of crossing over between the X and Y chromosomes. Oocytes from Ubc9K14R/K14R females show related but distinct defects in recombination and synapsis. Moreover, maintenance of the primordial follicle reserve is defective in Ubc9K14R/K14R females, and their fecundity is reduced. Finally, we identify the meiosis-specific homolog-axis protein SYCP3 as a direct target of UBC9 in vitro and show that SYCP3 modification is strongly stimulated by K14 auto-sumoylation. We infer that auto-sumoylation enables UBC9 to modify a subset of targets in vivo, helping to coordinate key events of meiotic prophase and enhance the survival of primordial follicles to maximize fecundity.

genetics↗

bioRxiv2026

A cohesion optimum underlies chromosome segregation fidelity in oocytes

Chromosome segregation is compromised in eggs from women of both early and advanced reproductive ages. Deteriorating cohesion causes premature separation of sister-chromatids in eggs from older females. We show that the converse is true for oocytes of adolescents, with excessive cohesion impeding segregation. Oocytes from juvenile mice show severe chromosome lagging in anaphase I, leading to nondisjunction or, in extreme cases, failure of the first meiotic division. These defects are suppressed by experimentally weakening cohesion or enhancing its resolution during anaphase I. By contrast, lagging and nondisjunction are rare in the oocytes of young adults because cohesion is inherently weaker. Thus, relative cohesion strength underlies both the frequency and type of segregation errors observed in eggs throughout the female reproductive lifespan. One-Sentence SummaryIn eggs, errors in chromosome segregation arise from age-dependent imbalances in how tightly chromosomes are held together.

cell biology↗

bioRxiv2026

Molecular architecture of meiotic pro-crossover factor HEI10 reveals coupling of higher-order assembly and ubiquitin chain formation

In meiosis, crossovers between homologous chromosomes generate genetic diversity and are required for accurate chromosome segregation, ensuring fertility. In mammals, HEI10 is one of three pro-crossover RING-domain factors implicated in protein modification by ubiquitin and/or SUMO and characterised by their dynamic accumulation at future crossover sites. However, the molecular architecture and enzymatic activity of mammalian HEI10 have remained unknown. Here, we show that human HEI10 has E3-ubiquitin ligase activity that depends on its higher-order assembly. We report the crystal structure of the HEI10 core, revealing how a 29-nm rod-like tetramer is formed through head-to-head association of two coiled-coil dimers that results in clustering of four RING domains around the molecular centre. HEI10 tetramers self-assemble through RING, coiled-coil, and C-terminal interfaces into fibrous and spherical higher-order structures. Structure-guided mutants show that higher-order assembly is required for HEI10 to catalyse K63-linked ubiquitin chain formation in vitro, with the most active species likely corresponding to a loose, non-fibrous network of assembled HEI10 molecules. Arabidopsis thaliana HEI10 retains the tetrameric core and higher-order assembly behaviour, suggesting a conserved principle of HEI10 function.

biochemistry↗

bioRxiv2026

Polycomb shapes active chromatin and promoter bivalency during ovarian reserve formation and activation

The ovarian reserve, a finite pool of long-lived non-growing oocytes established at birth, determines female reproductive lifespan, yet how these oocytes establish long-term quiescence while retaining the capacity for future growth and embryogenesis remains poorly understood. Here, we define a regulatory logic by which Polycomb repressive complexes shape stage-specific active chromatin remodeling during ovarian reserve formation and early oocyte growth. During ovarian reserve formation, H3K27ac, an active promoter- and enhancer-associated mark, undergoes extensive genome-wide redistribution. A key feature of this transition is CpG island promoter remodeling, in which many loci lose H3K27ac while gaining PRC1-dependent H2AK119ub, a repressive mark. This early reprogramming is followed during oocyte growth by acquisition of PRC2-dependent H3K27me3, de novo establishment of bivalent promoters, and protection of promoter regions from de novo DNA methylation. Oocyte growth is also accompanied by broad gains in both H3K27ac and H3K4me3, an active promoter-associated mark. Analyses of PRC1- and PRC2-deficient oocytes reveal unequal Polycomb contributions: PRC2 broadly constrains H3K27ac, whereas PRC1 more selectively shapes genome-wide H3K27ac redistribution and restricts H3K4me3 accumulation at bivalent promoters. Together, these findings identify staged active chromatin remodeling as an integral feature of perinatal oocyte development and reveal that Polycomb shapes chromatin state transitions as oocytes enter quiescence and become poised for future growth. One-Sentence SummaryPolycomb repressive complexes shape stage-specific active chromatin remodeling to establish quiescence and future promoter states during ovarian reserve formation and early oocyte growth.

developmental biology↗

bioRxiv2026

Cellector: A tool to detect foreign genotype cells in scRNAseq data with applications in leukemia and microchimerism.

The existence of rare, genetically distinct cells can occur in various samples such as transplant patients, naturally occurring microchimerism between maternal and fetal tissues, and cancer samples with sufficient mutational burden. Computational methods for detecting these foreign cells are vital to studying these biological conditions. An application that is of particular interest is that of leukemia patients post hematopoietic cell transplant (HCT). In many leukemias, a primary therapy is HCT, after which, the primary genotype of the bone marrow and blood cells should be of donor origin. If cells exist that are of the patients genotype and the cell type lineage of the particular leukemia, this is known as measurable residual disease (MRD). If the MRD is high enough, this may represent a relapse of the patients leukemia. Furthermore, accurately estimating the MRD is important for driving clinical decision making for these patients. Here we present Cellector, a computational method for identifying rare foreign genotype cells in single cell RNAseq (scRNAseq) datasets. We show cellector accurately detects microchimeric cells down to an exceedingly low percentage of these cells present (0.05% or lower).

bioinformatics↗

bioRxiv2026

SUMO modulates meiotic crossover rates between and within vertebrate species

Crossing over during meiosis drives genetic diversity and ensures the accurate segregation of homologous chromosomes. Variation in the rate of crossing over has been linked to evolutionary divergence and environmental adaptability, shaping fitness and responses to selective pressures. Despite its significance, the molecular mechanisms underlying this variation remain poorly understood. Crossover sites are selected from a large pool of potential sites initiated by programmed DNA double-strand breaks. Post-translational modification by SUMO (Small Ubiquitin-like Modifier) has been implicated in this process. Here, we show that crossover rate, chromosome length, and abundance of chromosome-associated SUMO are positively correlated across a range of vertebrate species, including mouse, chicken, pig, cattle, sheep, and goat. Crossover variation between goat breeds across the Indian subcontinent was also positively correlated with chromosomal SUMO level. Furthermore, modulating SUMO levels in cultured goat spermatocytes altered crossover frequency. Cumulatively, these observations point to a central role for SUMO in mediating crossover variation both between and within vertebrate species.

cell biology↗

bioRxiv2026

SUMO mediates the coordinate regulation of meiotic chromosome length and crossover rate

Meiotic prophase-I chromosomes are organized into linear arrays of chromatin loops anchored to proteinaceous axes that define the interaction interfaces for the pairing and synapsis of homologous chromosomes. Chromatin loop size and axial chromosome length are inversely correlated and vary widely both between and within species, including between the sexes. The molecular basis of this variation remains unclear. Here, we provide evidence that the small ubiquitin-like modifier, SUMO, regulates loop-axis organization in mouse meiosis. Our analysis shows that the longer axes of oocyte chromosomes contain more SUMO per unit length than the shorter axes of spermatocyte chromosomes. In mouse models, the loss of SUMO1 results in shorter axes and longer chromatin loops. Conversely, increased SUMO1 conjugation, caused by mutation of the SENP1 isopeptidase, produces longer axes with shorter loops. Axis length positively correlates with meiotic recombination. Accordingly, Sumo1 and Senp1 mutations respectively decrease and increase crossover frequency. These findings identify SUMO as a key regulator of meiotic chromosome architecture and suggest a molecular basis for the physiological variation in chromosome length and recombination rates seen among species, sexes, individuals, and individual meiocytes. GRAPHICAL ABSTRACT O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=101 SRC="FIGDIR/small/710713v1_ufig1.gif" ALT="Figure 1"> View larger version (31K): org.highwire.dtl.DTLVardef@145c465org.highwire.dtl.DTLVardef@160c8aborg.highwire.dtl.DTLVardef@1165b76org.highwire.dtl.DTLVardef@ced5e0_HPS_FORMAT_FIGEXP M_FIG C_FIG

cell biology↗

bioRxiv2025

HEIP1 orchestrates pro-crossover protein activity during mammalian meiosis

Meiotic crossovers are needed to produce genetically balanced gametes. In mammals, crossover formation is mediated by a conserved set of pro-crossover proteins via mechanisms that remain unclear. Here, we characterize a mammalian pro-crossover factor HEIP1. In mouse HEIP1 is essential for crossing over and fertility of both sexes. HEIP1 promotes crossing over by orchestrating the recruitment of other pro-crossover proteins, including the MutS{gamma} complex (MSH4- MSH5) and E3 ligases (HEI10, RNF212, and RNF212B), that are required to mature crossover sites and recruit the crossover-specific resolution complex MutL{gamma}. Moreover, HEIP1 directly interacts with HEI10, suggesting a direct role in controlling the recruitment of pro-crossover E3 ligases. During early stages of meiotic prophase I, HEIP1 interacts with the chromosome axes, independently of recombination, before relocalizing to the central region of the synaptonemal complex. We propose that HEIP1 is a new conserved master regulator of crossover proteins that controls different crossover maturation steps. Significance StatementCrossovers are essential to produce gametes by promoting the proper segregation of the homologous chromosomes. But, if misregulated, they can lead to genetic disorders, miscarriage and infertility. Their formation depends on the conserved pro-crossover factors, which repertoire is expanding and mode of action tightly regulated. This study highlights how the HEIP1 protein organizes pro-crossover protein activities in the mouse. Our findings show that HEIP1, by interacting early with chromosomes independently of recombination initiation, and orchestrating the recruitment of pro-crossover factors, including the MutSg complex and the RING proteins HEI10, RNF212 and RNF212B, is pivotal in this regulation. Our work is of significance to unravel crossover control during meiotic recombination, a conserved mechanism essential for gametes formation.

genetics↗

bioRxiv2025

Recruitment of Mre11 to recombination sites during meiosis

The Mre11 nuclease, part of the conserved MRX complex involved in the repair of DNA double-strand breaks (DSBs), is also essential to initiate meiotic recombination in budding yeast by promoting Spo11-induced DSBs. Recruitment of Mre11 to meiotic DSB sites depends on Rec114-Mei4 and Mer2 (RMM) that organize the meiotic DSB machinery by a mechanism involving biomolecular condensation. Here, we explored the role of Mre11 during meiosis and its relationship to RMM condensation. We show that both Mre11 and MRX complexes form DNA-dependent, hexanediol sensitive condensates in vitro. In vivo, Mre11 assembles into DNA damage-dependent foci in vegetative cells and DSB-independent foci in meiotic cells. In vitro condensates and in vivo foci both depend on the C-terminal intrinsically-disordered region (IDR) of Mre11. Importantly, while the Mre11 IDR is dispensable for vegetative DNA repair it is essential during meiosis. The C-terminus of Mre11 forms a short -helix that binds a conserved region of Mer2, and mutating residues within this interface reduces Mre11 foci and DSB formation. Finally, we identified a SUMO-interacting motif within the Mre11 IDR that enhances recruitment of Mre11 during meiosis and facilitates DSB formation. This work identifies multiple mechanisms that collaborate to recruit Mre11 during meiosis to initiate recombination.

molecular biology↗

bioRxiv2025

An Epigenomic Roadmap Primes Non-Growing Oocytes for Maturation and Early Embryogenesis

Female reproductive lifespan is defined by long-lived, non-growing oocytes (NGOs) that comprise the ovarian reserve. NGOs are assumed to acquire the epigenetic marks that will define the early embryo only after they exit the ovarian reserve and become activated for growth. Contrary to this dogma, we show that mouse NGOs possess abundant histone modifications that both underlie maintenance of the ovarian reserve and prime the epigenome of growing oocytes for early embryogenesis. As NGOs are established around birth, Polycomb Repressive Complex 1 (PRC1) mediates abundant H2AK119 ubiquitylation and reprograms the H3K27 acetylation landscape, which is essential to maintain the ovarian reserve. Importantly, the PRC1-driven epigenetic state of NGOs provides a blueprint for subsequent generation of a PRC2-catalyzed H3K27 trimethylation profile in growing oocytes that is characterized by broad domains and DNA methylation-independent imprints that are transmitted to the embryo. Thus, Polycomb complexes play pivotal roles in priming the NGO epigenome for oocyte maturation and early embryogenesis. HighlightsNon-growing oocytes have rich epigenetic landscapes to prime subsequent development Reprogramming of H3K27ac by PRC1 is required to maintain non-growing oocytes PRC1-catalyzed H2AK119ub provides a blueprint for PRC2-H3K27me3 in growing oocytes In non-growing oocytes, PRC1 primes DNA methylation-independent H3K27me3-imprinting

developmental biology↗

bioRxiv2024

Tethering ferredoxin-NADP+ reductase to photosystem I promotes photosynthetic cyclic electron transfer

Fixing CO2 via photosynthesis requires ATP and NADPH. Linear electron transfer (LET) supplies both metabolites, yet depending on environmental conditions, additional ATP is required which can be generated by cyclic electron transfer (CET). How the balance between LET and CET is set remains largely unknown. Ferredoxin(FD)-NADP+ reductase (FNR) has been suggested to act as the switch, channelling photosynthetic electrons to LET when it is bound to photosystem I (PSI) or CET when bound to cytochrome b6f. The essential role of FNR in LET precludes the use of a direct gene knock-out to test this hypothesis. We circumvented this problem by using CRISPR-Cas9 gene editing in Chlamydomonas reinhardtii to create a chimeric form of FNR tethered to PSI via PSAF. Chimeric FNR mutants exhibited impaired photosynthetic growth relative to the wild type, and decreased LET and non-photochemical quenching (NPQ) under high CO2 conditions. However, higher rates of CET and increased NPQ were found in the mutants under low CO2 or low O2 conditions where CET is normally up-regulated. Thus, rather than PSI-FNR interactions promoting LET, tethering FNR to PSI promotes CET at the expense of LET and CO2 fixation.

biochemistry↗

bioRxiv2024

Synaptonemal complex protects double-Holliday junctions during meiosis

Chromosomal linkages formed through crossover recombination are essential for accurate segregation of homologous chromosomes during meiosis1. The DNA events of recombination are linked to structural components of meiotic chromosomes2. Imperatively, the biased resolution of double-Holliday junction intermediates (dHJs) into crossovers3,4 occurs within the synaptonemal complex (SC), the meiosis-specific structure that mediates end-to-end synapsis of homologs during the pachytene stage5,6. However, the SCs roles in crossover-specific dHJ resolution remains unclear. Here, we show that key SC components function through dependent and interdependent relationships to protect dHJs from aberrant "dissolution" into noncrossover products. Conditional ablation experiments reveal that cohesin, the core of SC lateral elements, is required to maintain both synapsis and dHJ-associated crossover recombination complexes (CRCs) during pachytene. The SC central region transverse-filament protein is also required to maintain CRCs. Reciprocally, stability of the SC central region requires the continuous presence of CRCs, thereby coupling synapsis and desynapsis to dHJ formation and resolution. However, dHJ protection and maintenance of CRCs can occur without end-to-end homolog synapsis mediated by the central element of the SC central region. We conclude that local ensembles of SC components are sufficient to enable crossover-specific dHJ resolution and thereby ensure the linkage and segregation of homologous chromosomes.

cell biology↗

bioRxiv2023

Distinct and interdependent functions of three RING proteins regulate recombination during mammalian meiosis

During meiosis, each pair of homologous chromosomes becomes connected by at least one crossover, as required for accurate segregation at the first division, and adjacent crossovers are widely separated thereby limiting total numbers. In coarsening models, crossover patterning results from nascent recombination sites competing to accrue a limiting pro-crossover RING-domain protein (COR) that diffuses between synapsed chromosomes. Here we delineate the localization dynamics of three mammalian CORs in mouse and determine their interdependencies. RNF212, HEI10 and a new member RNF212B show divergent spatiotemporal dynamics along synapsed chromosomes, including profound differences in spermatocytes and oocytes, that are not easily reconciled by elementary coarsening models. Contrasting mutant phenotypes and genetic requirements indicate that RNF212B, RNF212, and HEI10 play distinct but interdependent functions in regulating meiotic recombination and coordinating the events of meiotic prophase-I by integrating signals from DNA breaks, homolog synapsis, the cell-cycle, and incipient crossover sites. SIGNIFICANCEMeiosis produces haploid gametes by precisely halving the chromosome complement. Crossing over between homologous chromosomes (homologs) is essential for their accurate segregation and defects are associated with infertility, miscarriage, and congenital disease. Factors that ensure crossing over between each pair of homologs include mammalian RING-domain proteins RNF212, HEI10, and RNF212B, alleles of which are linked to infertility and heritable variation in crossover rate. This study focuses on understanding the functions and relationships between pro-crossover RING proteins (CORs) in mouse, providing important insights into their roles in regulating recombination, the DNA repair process that gives rise to crossovers. Notably, chromosomal localization dynamics of the three CORs are distinct and show striking sexual dimorphism with important implications for models of crossover control.

genetics↗

bioRxiv2023

Increasing equity in science requires better ethics training: a course by trainees, for trainees

Despite the profound impacts of scientific research, few scientists have received the necessary training to productively discuss the ethical and societal implications of their work. To address this critical gap, we - a group of predominantly human genetics trainees - developed a course on genetics, ethics, and society. We intend for this course to serve as a template for other institutions and scientific disciplines. Our curriculum positions human genetics within its historical and societal context and encourages students to evaluate how societal norms and structures impact the conduct of scientific research. We demonstrate the utility of this course via surveys of enrolled students and provide resources and strategies for others hoping to teach a similar course. We conclude by arguing that if we are to work towards rectifying the inequities and injustices produced by our field, we must first learn to view our own research as impacting and being impacted by society.

genetics↗

bioRxiv2023

Transcriptomics and chromatin accessibility in multiple African population samples

Mapping the functional human genome and impact of genetic variants is often limited to European-descendent population samples. To aid in overcoming this limitation, we measured gene expression using RNA sequencing in lymphoblastoid cell lines (LCLs) from 599 individuals from six African populations to identify novel transcripts including those not represented in the hg38 reference genome. We used whole genomes from the 1000 Genomes Project and 164 Maasai individuals to identify 8,881 expression and 6,949 splicing quantitative trait loci (eQTLs/sQTLs), and 2,611 structural variants associated with gene expression (SV-eQTLs). We further profiled chromatin accessibility using ATAC-Seq in a subset of 100 representative individuals, to identity chromatin accessibility quantitative trait loci (caQTLs) and allele-specific chromatin accessibility, and provide predictions for the functional effect of 78.9 million variants on chromatin accessibility. Using this map of eQTLs and caQTLs we fine-mapped GWAS signals for a range of complex diseases. Combined, this work expands global functional genomic data to identify novel transcripts, functional elements and variants, understand population genetic history of molecular quantitative trait loci, and further resolve the genetic basis of multiple human traits and disease.

genomics↗

bioRxiv2022

Reduced oocyte quality and exacerbation of the maternal age effect are enduring consequences of low-level atrazine exposure in mouse

BACKGROUNDEgg development has unique features that render it vulnerable to environmental perturbation. The herbicide atrazine is an endocrine disruptor shown to have detrimental effects on reproduction across a number of vertebrate species. OBJECTIVESTo determine whether exposure to low levels of atrazine impairs meiosis in female mammals using a mouse model; in particular, whether and how the fidelity of oocyte chromosome segregation is affected, and whether the aging-related aneuploidy is exacerbated. METHODSFemale C57BL/6J mice were exposed to two levels of atrazine in drinking water, with the lower level corresponding to detected environmental contamination. To model exposure during development, atrazine was ingested by pregnant females at 0.5 days post coitum and continued until pups were weaned at 21 days post-partum. For adult exposure, 2-month-old females ingested atrazine for 3 months. For each exposure group, various indicators of oocyte quality were determined, including developmental capacity and chromosomal abnormalities during the two meiotic divisions. RESULTSDevelopmental exposure caused only minor effects on the fetal events of meiotic prophase-I and establishment of initial follicle pools. However, ovulation was enhanced while oocyte quality was significantly reduced. At the chromosome level, misalignment and numerical and structural abnormalities were increased at both meiotic divisions. Furthermore, fertilization efficiency was impaired in vitro, and apoptosis was elevated in blastocysts derived from the eggs of atrazine-exposed females. Similar levels of chromosomal defects were seen in oocytes following both developmental and adult exposure regimens suggesting that quiescent primordial follicles may be the consequential targets of atrazine. Importantly, defects were observed long after exposure was terminated. Moreover, dramatic increases in chromosomally abnormal oocytes were seen in older mice indicating that atrazine exposure during development exacerbates the effects of maternal aging on oocyte quality. Indeed, analogous to the effects of maternal age, atrazine exposure resulted in weakened cohesion between sister chromatids. CONCLUSIONLow-level atrazine exposure causes persistent changes to the female mammalian germline with potential consequences for reproductive lifespan and congenital disease.

pharmacology and toxicology↗

bioRxiv2020

PCNA activates the MutLγ endonuclease to promote meiotic crossing over

During meiosis, crossover recombination connects homologous chromosomes to direct their accurate segregation1. Defects in crossing over cause infertility, miscarriage and congenital disease. Accordingly, each pair of chromosomes attains at least one crossover through processes that designate and then implement crossing over with high efficiency2. At the DNA level, crossing over is implemented through the formation and biased resolution of double-Holliday Junction intermediates3-5. A central tenet of crossover resolution is that the two Holliday junctions are resolved in opposite planes by targeting nuclease incisions to specific DNA strands6. Although the endonuclease activity of the MutL{gamma} complex has been implicated in crossover-biased resolution7-12, the mechanisms that activate and direct strand-specific cleavage remain unknown. Here we show that the sliding clamp, PCNA, is important for crossover-biased resolution. In vitro assays with human enzymes show that hPCNA and its loader hRFC are sufficient to activate the hMutL{gamma} endonuclease under physiological conditions. In this context, the hMutL{gamma} endonuclease is further stimulated by a co-dependent activity of the pro-crossover factors hEXO1 and hMutS{gamma}, the latter of which binds Holliday junctions13. hMutL{gamma} also specifically binds a variety of branched DNAs, including Holliday junctions, but canonical resolvase activity is not observed implying that the endonuclease incises adjacent to junction branch points to effect resolution. In vivo, we show that budding yeast RFC facilitates MutL{gamma}-dependent crossing over. Furthermore, PCNA localizes to prospective crossover sites along synapsed chromosomes. These data highlight similarities between crossover-resolution and DNA mismatch repair14-16 and evoke a novel model for crossover-specific dHJ resolution during meiosis.

biochemistry↗

bioRxiv2019

Delineation of the SUMO-Modified Proteome Reveals Regulatory Functions Throughout Meiosis

Protein modification by SUMO helps orchestrate the elaborate events of meiosis to faithfully produce haploid gametes. To date, only a handful of meiotic SUMO targets have been identified. Here we delineate a multidimensional SUMO-modified meiotic proteome in budding yeast, identifying 2747 conjugation sites in 775 targets, and defining their relative levels and dynamics. Modified sites cluster in disordered regions and only a minority match consensus motifs. Target identities and modification dynamics imply that SUMOylation regulates all levels of chromosome organization and each step of homologous recombination. Execution-point analysis confirms these inferences, revealing functions for SUMO in S-phase, the initiation of recombination, chromosome synapsis and crossing over. K15-linked SUMO chains become prominent as chromosomes synapse and recombine, consistent with roles in these processes. SUMO also modifies ubiquitin, forming hybrid oligomers with potential to modulate ubiquitin signaling. We conclude that SUMO plays diverse and unanticipated roles in regulating meiotic chromosome metabolism.

genetics↗