Biology subjects

Hebbar, P.

Publications and source records attributed to Hebbar, P..

16 recordsLinked to original sources

bioRxiv2026

Pangenome alignment reveals global diversity and evolution of human centromeric regions

Centromeres play essential roles in chromosome segregation and genome stability, yet they remain among the least characterized regions of the human genome. Despite advances in long-read sequencing and complete genome assembly, the extreme repetitiveness and structural complexity of these regions still challenge population-scale analysis, obscuring their mutational dynamics. The Human Pangenome Reference Consortium has now accurately assembled over 6,000 centromeres, providing an opportunity to catalog global centromere variation. However, centromeric regions have been systematically excluded from pangenome alignments due to the technical challenge of aligning their highly repetitive tandem arrays and extreme structural variability. Here we introduce Centrolign, a graph-based multiple sequence alignment tool that combines a uniqueness-driven objective function with partial-order partial-order alignment to accurately align alpha satellite higher-order repeats. By prioritizing rare matches within tandem arrays and leveraging extended centromere-spanning haplotypes formed by suppressed recombination, Centrolign produces progressive multiple sequence alignments that preserve ancestral repeat organization. Applied across human centromeres, these alignments reveal the phylogenetic structure of similar satellite array haplotypes and enable precise estimation of variation rates, structural variant frequencies, and spatial patterns of mutation within satellite arrays. Integrating Centrolign graphs with repeat annotation tools and pangenome mapping algorithms allows accurate variant calling and genotyping from long reads without prior assembly. Moreover, we show that centromere haplotypes can be accurately subtyped with k-mers alone. Together, these advances establish a robust framework for incorporating centromeres into broader pangenomes, and population genomics in general, advancing our understanding of human genome evolution and diversity.

genomics↗

bioRxiv2026

HPRC2: A human pangenome reference with near-complete coverage of common genetic variation

A pangenome reference overcomes the inherent limitation of any individual reference genome by integrating the variation present in a population. We present the Human Pangenome Reference Consortiums (HPRC) Release 2 (HPRC2), an openly available, second phase pangenome that is an approximately fivefold expansion in genome number over HPRC Release 1 (HPRC1) and measurable improvement in genome completeness, contiguity, and accuracy. Selecting samples with a principled algorithm prioritising common variant coverage, HPRC2 contributes 460 haplotypes that together capture over 99% of common variation observed in the All of Us Research Program v8 cohort. Combining high-coverage long and ultra-long reads with modern assemblers and polishers, we produce thousands of telomere-to-telomere (T2T) chromosomes, and relative to HPRC1 halve the number of structurally unreliable regions as well as individual base errors per haplotype. We complement the assemblies with whole genome multiple alignments and gene annotations, and derive formal pangenome coordinate systems for addressing off-reference variation, demonstrating that individual human genomes contain more than one hundred thousand variants not succinctly described with respect to existing reference genomes. We also present the first matched long-read backed pantranscriptome and panepigenome at this scale, provide continuous local-ancestry estimates spanning every genome, and outline a host of new tools and applications that leverage the pangenome resource for improved genomics analysis.

genomics↗

bioRxiv2026

A Pan-pangenome illuminates complex structural variation and selection in humans, chimpanzees, and bonobos

Complete, haplotype-resolved genome assemblies have provided unprecedented insight into the evolution of structurally complex, rapidly evolving regions of human genomes1-6; however, population-scale pangenome resources of our closest relatives, chimpanzees and bonobos (genus, Pan), are necessary to ascertain the origins and evolutionary context of these loci. Here, we sequence and assemble 58 haplotypes from four distinct Pan clades to high contiguity (median contig NG50=54 Mb), including eight near-T2T genomes. These genomes reveal previously intractable genetic variation increasing estimates of genome-wide genetic diversity 6-37% across populations compared to short-read estimates7. We identify recurrent structural polymorphisms across species impacting genes associated with immune response and host-pathogen interaction and find that structural variants (SVs) are 170- to 260-fold more likely than single nucleotide variants (SNVs) to exhibit high-impact effects across species. Contrasting SV patterns across primates we find that transposable element mutation rates differ by as much as threefold between species. We show that human disease-associated short tandem repeat (TR) loci have uniquely expanded in humans sensitizing our species to these TR-expansion disorders. Physically phased haplotypes enable reconstruction of genome-wide genealogical histories, uncovering ancient, functional genetic variation maintained by balancing selection, as well as signatures of recent adaptation in chimpanzee subspecies. Several malaria-associated loci exhibit ancient structural polymorphism, including the African great ape-specific glycophorin (GYP) gene expansion. We characterize the sequence, structure, and composition of diverse glycophorin haplotypes in humans and chimpanzees. We identify independent malaria-protective GYPA-B fusion events in humans and novel chimpanzee glycophorin genes resulting from both ancient and recent fusion events demonstrating parallel adaptations to pathogen resistance across hominins. Together, our resource highlights the critical importance of nonhuman primate population-scale pangenomics for understanding the evolution of complex genome structures and the biodiversity of our endangered closest living relatives.

evolutionary biology↗

bioRxiv2026

Population-scale Y chromosome assemblies reveal recurrent remodeling within constrained architectures

The human Y chromosome is among the most structurally dynamic chromosomes in the human genome, yet much of its diversity remains unresolved because of extensive palindromes, ampliconic gene families, satellite-rich heterochromatin and large segmental duplications. What remained unclear was how these diverse forms of variation fit together across the full chromosome, how often similar structures recur in different lineages, and which aspects of organization remain constrained despite rapid sequence turnover. Here, we generated and analyzed 142 nearly complete human Y chromosome assemblies from 17 major haplogroups spanning approximately 180,000 years of evolution, creating a population-scale resource for studying Y chromosome biology and diversity. These assemblies show that structural change on the Y chromosome is recurrent but constrained, even in its most repetitive regions. In the fertility-associated azoospermia factor c (AZFc) region, recurrent inversions, deletions, and complex rearrangements generate a limited repertoire of structural haplotypes. Multicopy ampliconic gene families follow distinct evolutionary paths: DAZ paralogues differ in structural constraint, RBMY evolves within a modular array, and TSPY copy number varies mainly through local expansion and contraction. The centromere and Yq12 heterochromatin vary greatly in size but retain a stable higher-order organization, including a single hypomethylated centromeric core and conserved Yq12 repeat composition and orientation. Methylation across palindromic and ampliconic regions is likewise structured by repeat class, copy order and local architecture. Together, these results provide a population-scale resource for the human Y chromosome and show that its rapid structural evolution is repeatedly funneled into a limited set of architectural outcomes.

genomics↗

bioRxiv2026

The diploid reference genome of a human embryonic stem cell line

Advances in DNA sequencing and assembly technologies are spurring a shift from haploid reference genomes to sample-specific diploid assemblies. Here, we generated the first telomere-to-telomere (T2T) diploid reference for the widely used human embryonic stem cell (hESC) line, H9 (WAe009-A). This haplotype-resolved assembly is highly accurate with comprehensive annotation of genes, segmental duplications, methylation, and chromatin conformation. Pangenomic and phased-locus inference point to H9s mixed ancestry with a predominant European component. H9-specific genomic features include near-perfect telomeres [~]1.65-fold longer than other T2T assemblies, consistent with telomerase activity during pluripotency; chromosome 17 inversions that can predispose offspring to neurological syndromes; and expansions of ncRNA clusters, with overall genomic stability maintained despite extensive culturing. Mapping multi-omic datasets to the genome, we demonstrate the power of this resource for allele-specific, high-precision transcriptomic, genetic, and epigenetic analyses, with far-reaching implications for human development and disease.

genomics↗

bioRxiv2026

A Complete Genome for the Common Marmoset

The common marmoset is a New World monkey (NWM) commonly used as a model organism to investigate questions in primate evolution and human disease, including Alzheimers and other neurodegenerative diseases, as well as neuropsychiatric disorders. Here we present the first telomere-to-telomere (T2T) reference genome for the common marmoset, adding over 88 Mb of sequence and resolving challenging genomic regions. An additional near-T2T assembly from a second unrelated individual yields a total of four high-quality haplotypes for analysis. The improved contiguity and accuracy of these assemblies enable unprecedented insights into complex and rapidly evolving genomic regions such as centromeres, sex chromosomes, ribosomal DNA (rDNA) structure, and the major histocompatibility complex (MHC). We fully resolved all marmoset centromeres, uncovering dimeric alpha satellites with chromosomal specificity and stratified inactive layers documenting ancestral centromere turnover. We assembled six acrocentric autosomes with gene-poor, satellite-rich short arms and provide evidence that most of them can harbor rDNA and all of them share large pseudo-homologous regions (PHRs). The Y chromosome, but not the X chromosome, carries active rDNA and PHRs, and the rDNA copy number is sexually dimorphic. Chromosomes that share PHRs also share closely related centromeric satellite DNA, supporting a model of ongoing recombinational exchange between heterologous chromosomes facilitated by rDNA. We discovered multiple novel, marmoset-specific MHC genes that are predicted to protect against pathogens encountered in its environment. Leveraging this complete reference, we further identified over 500 transcribed genes with transcript models or expansions specific to the marmoset lineage. Together with additional long-read marmoset assemblies, these genomes were used to construct a marmoset pangenome, providing a robust reference framework for short-read mapping across diverse individuals. This resource will improve the utility of the common marmoset as a biomedical model organism and fill key gaps in our understanding of primate evolution.

genomics↗

bioRxiv2026

The complete genome of the KOLF2.1J reference iPSC line

While induced pluripotent stem cells (iPSCs) have gained popularity in studying neurodegenerative diseases, the heterogeneity of stem cells used across studies impacts cross-study comparison. The iPSC Neurodegenerative Disease Initiative (iNDI) selected the KOLF2.1J cell line and prioritized its use as a reference standard for studying the effects of pathogenic variants on cell biology due to its stability and neutral neurodegenerative disease genetic risk. This cell line, and its derivatives expressing over 100 variants related to Alzheimers disease, Parkinsons disease, and other neurological diseases, are available for academic and industry access. Current genomic data analyses are limited by the use of a human reference genome that does not capture the complete genetic background of a given iPSC line. While in the future this issue may be partially mitigated by the creation of a comprehensive human pangenome, previous work has shown that generating custom genomes is of value both to characterize the variation present and to serve as a more appropriate genomic reference. Here, we generated and characterized a custom complete genome assembly from KOLF2.1J. Mapping of sequencing reads to a personalized diploid assembly results in more comprehensive mapping compared to traditional linear references (i.e GRCh38). In addition, we provide a comprehensive custom gene annotation along with isoform expression and differential methylation analyses across multiple cell types. The assembly and all additional data is browsable and publicly available. This resource will enable more accurate investigation of the KOLF2.1J cell line and any genomics data generated compared to using traditional generalized references, while also serving as a foundational approach for establishing custom reference assemblies for other high-value iPSC lines.

genomics↗

bioRxiv2026

Evaluating genome assemblies with HMM-Flagger

HMM-Flagger is a reference-free tool for detecting structural errors in haplotype-resolved genome assemblies based upon the coverage of mapped reads. It models read coverage with a hidden Markov model augmented by a Gaussian autoregressive process, which enables classifying coverage anomalies as erroneous blocks, false duplications, or collapsed blocks. Trained and tested on synthetic misassemblies, it detected synthetic errors using Pacific Biosciences HiFi and Oxford Nanopore Technologies R10 data with F1 scores of 78.4% and 60.4% respectively. When applied to six HG002 assemblies it revealed multiple large misassemblies including false duplications and collapse events in human satellites. Applied to assemblies from the Human Pangenome Reference Consortium (HPRC), HMM-Flagger demonstrated substantial improvements from release 1 (0.94% error rate) to release 2 (0.38%), reflecting technological advances. HMM-Flagger also validated NOTCH2NL assemblies in HPRC release 2 and confirmed the correctness of three novel structural configurations.

bioinformatics↗

bioRxiv2025

Origin and evolution of acrocentric chromosomes in human and great apes

The short arms of human acrocentric chromosomes are characterized by nucleolar organizer regions essential for ribosome biogenesis, but their highly repetitive nature has hindered genomic analysis. Leveraging the recently completed genomes of all major ape lineages, we identified recurrent features of their acrocentrics, including enriched repeat classes, centromere repositioning by whole-arm inversion, interchromosomal sequence exchange, and birth-and-death evolution of multiple gene families. Together, these processes have enabled the repeated amplification and diversification of the FRG1 gene family over 25 million years of ape evolution, and, in gorilla, the formation and amplification of a novel IGSF3-GGT fusion gene under positive selection. Similar evolutionary events also explain the distribution of segmental duplications and heterochromatin in the modern human genome, predisposing it to karyotypic abnormalities such as Robertsonian translocations. Our findings highlight acrocentric chromosomes as key drivers of evolution in the great apes, with implications for speciation, adaptation, and clinical genomics.

genomics↗

bioRxiv2025

Complete genomes of a multi-generational pedigree to expand studies of genetic and epigenetic inheritance

Pedigree analysis remains the gold standard for rare disease diagnostics, yet whole genome sequencing studies typically omit critical regions like centromeres, telomeres, and acrocentric chromosome p-arms. Here, we present telomere-to-telomere (T2T) reference genomes for four self-identified African American individuals of admixed ancestry spanning three generations. Our parent-of-origin assigned, chromosome-level assemblies revealed precise meiotic recombination breakpoints in previously inaccessible regions, including recombination events across acrocentric and subtelomeric sequences. Centromeric regions were highly stable, with multi-megabase arrays inherited intact across three generations, while the position of kinetochore assembly sites remained consistent and predominantly associated with the p-arm proximal region. The relative lengths of telomeres on individual chromosomes were maintained across generations. Using a targeted rDNA assembly approach, we reconstructed a complete megabase-scale ribosomal DNA (rDNA) array corresponding to the paternal chromosome 14. This openly available pedigree provides a benchmark dataset for studying recombination and genetic and epigenetic variation across the complete genome.

genomics↗

bioRxiv2025

Genetic diversity and regulatory features of human-specific NOTCH2NL duplications

NOTCH2NL (NOTCH2-N-terminus-like) genes arose from ape-specific chromosome 1 segmental duplications implicated in human brain cortical expansion, including an incomplete NOTCH2 gene. Genetic characterization of these loci and their regulation is complicated because they are embedded in large, nearly identical duplications that predispose to recurrent microdeletion syndromes. Using near-complete long-read assemblies generated from 70 human and 12 ape haploid genomes, we show independent recurrent duplication among apes with protein-coding copies emerging in humans 2.2-3.7 million years ago. We distinguish NOTCH2NL paralogs present in every human haplotype (NOTCH2NLA) from copy number variable ones. We also characterize large-scale structural variation, including gene conversion, for 28% of haplotypes leading to a previously undescribed paralog, NOTCH2tv. Finally, we apply Fiber-seq and long-read transcript sequencing to human dorsal forebrain organoids to characterize the regulatory landscape and find that the most fixed paralogs, NOTCH2 and NOTCH2NLA, harbor the greatest number of paralog-specific elements potentially driving their regulation.

genomics↗

bioRxiv2024

Highly accurate assembly polishing with DeepPolisher

Accurate genome assemblies are essential for biological research, but even the highest quality assemblies retain errors caused by the technologies used to construct them. Base-level errors are typically fixed with an additional polishing step that uses reads aligned to the draft assembly to identify necessary edits. However, current methods struggle to find a balance between over- and under-polishing. Here, we present an encoder-only transformer model for assembly polishing called DeepPolisher, which predicts corrections to the underlying sequence using Pacbio HiFi read alignments to a diploid assembly. Our pipeline introduces a method, PHARAOH (Phasing Reads in Areas Of Homozygosity), which uses ultra-long ONT data to ensure alignments are accurately phased and to correctly introduce heterozygous edits in falsely homozygous regions. We demonstrate that the DeepPolisher pipeline can reduce assembly errors by half, with a greater than 70% reduction in indel errors. We have applied our DeepPolisher-based pipeline to 180 assemblies from the next Human Pangenome Reference Consortium (HPRC) data release, producing an average predicted Quality Value (QV) improvement of 3.4 (54% error reduction) for the majority of the genome.

bioinformatics↗

bioRxiv2024

Complete sequencing of ape genomes

We present haplotype-resolved reference genomes and comparative analyses of six ape species, namely: chimpanzee, bonobo, gorilla, Bornean orangutan, Sumatran orangutan, and siamang. We achieve chromosome-level contiguity with unparalleled sequence accuracy (<1 error in 500,000 base pairs), completely sequencing 215 gapless chromosomes telomere-to-telomere. We resolve challenging regions, such as the major histocompatibility complex and immunoglobulin loci, providing more in-depth evolutionary insights. Comparative analyses, including human, allow us to investigate the evolution and diversity of regions previously uncharacterized or incompletely studied without bias from mapping to the human reference. This includes newly minted gene families within lineage-specific segmental duplications, centromeric DNA, acrocentric chromosomes, and subterminal heterochromatin. This resource should serve as a definitive baseline for all future evolutionary studies of humans and our closest living ape relatives.

genomics↗

bioRxiv2023

The Complete Sequence and Comparative Analysis of Ape Sex Chromosomes

Apes possess two sex chromosomes--the male-specific Y and the X shared by males and females. The Y chromosome is crucial for male reproduction, with deletions linked to infertility1. The X chromosome carries genes vital for reproduction and cognition2. Variation in mating patterns and brain function among great apes suggests corresponding differences in their sex chromosomes. However, due to their highly repetitive nature and incomplete reference assemblies, ape sex chromosomes have been challenging to study. Here, using the methodology developed for the telomere-to-telomere (T2T) human genome, we produced gapless assemblies of the X and Y chromosomes for five great apes (chimpanzee, bonobo, gorilla, Bornean and Sumatran orangutans) and a lesser ape, the siamang gibbon. These assemblies allowed us to untangle the intricacies of ape sex chromosome evolution. We found that, compared to the Xs, the ape Ys vary greatly in size and have low alignability and high levels of structural rearrangements. This divergence on the Y arises from the accumulation of lineage-specific ampliconic regions, palindromes, transposable elements, and satellites. Our analysis of Y chromosome genes revealed expansions of multi-copy gene families and signatures of purifying selection. Thus, the Y exhibits dynamic evolution, while the X is more stable. Mapping short-read sequencing data to these assemblies revealed diversity and selection patterns on sex chromosomes of >100 great ape individuals. These reference assemblies are expected to inform human evolution and conservation genetics of nonhuman apes, all of which are endangered species.

genomics↗

bioRxiv2023

ANGPTL8 R59W variant influences inflammation through modulating NF-κB pathway under TNFα stimulation

BackgroundANGPTL8 is known to regulate lipid metabolism and inflammation. It interacts with ANGPTL3 and ANGPTL4 to regulate LPL activity, and with IKK/{beta} to modulate NF-{kappa}B activity. Further, a SNP leading to ANGPTL8 R59W variant associates with reduced LDL/HDL and increased FBG in Hispanic and Arab individuals, respectively. In this study, we investigate the impact of R59W variant on the inflammatory activity of ANGPTL8. MethodsANGPTL8 R59W variant was genotyped in a discovery cohort of 867 Arab individuals from Kuwait. Plasma levels of ANGPTL8 and inflammatory markers were measured and tested for associations with the genotype; the associations were tested for replication in an independent cohort of 278 Arab individuals. Impact of the ANGPTL8 R59W variant on NF-{kappa}B activity was examined using approaches including overexpression, luciferase assay, and structural modeling of binding dynamics. ResultsThe ANGPTL8 R59W variant was associated with increased circulatory levels of TNF and IL7. NF-{kappa}B activity, as assessed by the increased in the phosphorylation of IKK-/{beta} protein, I{kappa}B, and NF-{kappa}B p-65 in R59W variant compared to wild type, and TNF stimulation further elevated it. This finding was substantiated by increased luciferase activity of NF-{kappa}B p65 with the R59W variant. Modeled structural and binding variation due to R59W change in ANGPTL8 agreed with the observed increase in NF-{kappa}B activity. ConclusionANGPTL8 R59W is associated with increased circulatory TNF, IL7 and NF-{kappa}B p65 activity. Weak transient binding of ANGPTL8 R59W variant explains its regulatory role on the NF-{kappa}B pathway and inflammation.

biochemistry↗

bioRxiv2020

Fine-scale population structure reveals high genetic heterogeneity of the Kuwaiti population in the Arabian Peninsula

Recent studies have showed the diverse genetic architecture of the highly consanguineous populations inhabiting the Arabian Peninsula. Consanguinity coupled with heterogeneity is complex and makes it difficult to understand the bases of population-specific genetic diseases in the region. Therefore, comprehensive genetic characterization of the populations at the finest scale is warranted. Here, we revisit the genetic structure of the Kuwait population by analyzing genome-wide single nucleotide polymorphisms data from 583 Kuwaiti individuals sorted into three subgroups. We envisage a diverse demographic genetic history among the three subgroups based on drift and allelic sharing with modern and ancient individuals. Furthermore, our comprehensive haplotype-based analyses disclose a high genetic heterogeneity among the Kuwaiti populations. We infer the major sources of ancestry within the newly defined groups; one with an obvious predominance of sub-Saharan/Western Africa mostly comprising Kuwait-B individuals, and other with West Eurasia including Kuwait-P and Kuwait-S individuals. Overall, our results recapitulate the historical population movements and reaffirm the genetic imprints of the legacy of continental trading in the region. Such deciphering of fine-scale population structure and their regional genetic heterogeneity would provide clues to the uncharted areas of disease-gene discovery and related associations in populations inhabiting the Arabian Peninsula.

genomics↗