Biology subjects

Karthikeyan, S.

Publications and source records attributed to Karthikeyan, S..

15 recordsLinked to original sources

bioRxiv2026

Sustained epigenetic rejuvenation of serially engrafting human iPSC-derived HSCs

Hematopoietic stem cell (HSC) function declines with age, contributing to immunosenescence and inferior transplantation outcomes. Here, we generated iPSC-derived HSCs (iHSCs) from multiple adult donors and performed integrated epigenetic, transcriptional, telomeric, and functional analyses to see if they retain youthful identity across differentiation and serial transplantation. Longitudinal DNA methylation profiling revealed that, independent of donor age, epigenetic age was reset to near zero in iPSCs and remained under seven years across differentiation and transplantation. In contrast, hematopoietic identity was established through a two-phase process: directional remodeling during in vitro differentiation extinguished pluripotency programs and initiated hematopoietic regulatory networks, while long-term engraftment was associated with a second wave of promoter methylation differences that converged toward primary adult HSCs. Notably, methylation at age-associated sites and global entropy remained stable across both phases, and single-cell telomere analysis demonstrated restoration of telomere length in iHSCs compared to primary adult HSCs. Youthful epigenetic features were maintained through secondary transplantation. These findings demonstrate that long-term HSC identity can be achieved independently of epigenetic aging and establish a framework for evaluating rejuvenated stem cell-derived grafts in regenerative medicine.

cell biology↗

bioRxiv2026

WNT11 suppresses tumor initiation and invasion by inactivating RAC1

WNT11, a non-canonical WNT ligand, plays well-defined roles in development and tissue architecture; however, its function in cancer remains ambiguous. Here, we characterize WNT11 as a context-dependent suppressor of cancer stemness, invasion, and in vivo tumor formation in human epithelial cancer models. We show that WNT11 upregulation reduces the expression of stemness-promoting genes, suppresses epithelial-to-mesenchymal transition, and inhibits sphere formation and tumor growth. Conversely, downregulation of WNT11 enhances these aggressive malignant properties of cancer cells. Mechanistically, we found that the ability of WNT11 to inhibit RAC1 GTPase activation is essential for its regulation of invasion and self-renewal. In cells unresponsive to WNT11, the connectivity between WNT11 and RAC1 activity is disengaged. Direct manipulation of RAC1 activity in these cells recapitulates the phenotype and molecular signature of WNT11-responsive cells, establishing RAC1 as a critical effector of WNT11-mediated tumor suppression. Taken together, these findings identify the cellular context in which WNT11 suppresses RAC1 activation as a key determinant of its anti-tumor effects and provide a mechanistic framework for understanding the diverse, and sometimes opposing, roles of WNT11 reported in cancer.

cancer biology↗

bioRxiv2026

SIP-enabled multi-omics reveals soil microbiome responses to drought and rehydration

The activity of the soil microbiome, and its balance of anabolic (organic C consuming) and catabolic (CO2-releasing) reactions, determines the magnitude and direction of soil carbon fluxes. Over half a century of research has revealed that soil water dynamics are key controllers of microbial activity. With increasing hydroclimate volatility expected across many regions of the Earth, there is a greater need to describe and quantify microbial responses to drought and rehydration cycles. In this study, we conducted rainfall exclusion experiments at two archetypical Mediterranean-type field sites. After rainfall exclusion and subsequent soil rehydration, we applied a SIP-enabled, multi-omics methodology to generate a multi-faceted case study of microbial growth, greenhouse gas fluxes, and the forms of carbon that drive both. Our results indicate that rehydration increases microbial anabolic processes by orders of magnitude, shifting cell generation times from years to days within just minutes. High-intensity drought increases the lag period before microbial growth resumes, but both stable-isotope probing and metagenomic inference agree that microbial communities exhibit greater capacity for rapid growth following drought stress. Furthermore, significant shifts in the soil metabolome are observed following drought and rehydration, implicating specific osmolytes as key to the microbial response and indicating metabolite diversity as a key modulator of microbiome functioning. Together, our results provide constraints on microbial activity rates in soil and mechanisms underpinning microbial responses to drought and rewetting. These findings motivate further research into microbial responses under increasingly volatile hydroclimate regimes and downstream contributions to the global carbon cycle. Significance StatementSoil is a major global store and source of carbon. The microbiome determine the fate of soil organic carbon, and the microbiome is ultimately controlled by soil water dynamics. Early, innovative experiments by H.F. Birch defined "The Birch Effect" - the observation that soils emit CO2 following drying and subsequent rehydration. However, it remains unclear when, and to what magnitude, soil microorganisms are actively growing following this rehydration, and what biological mechanisms explain the observed CO2 pulse. In this work, we apply an array of methodologies to address this question, describing rates of microbial growth during drought and rewetting. Our results provide crucial insights into how soil microbiomes will respond to increasing hydroclimate volatility across the globe.

microbiology↗

bioRxiv2026

Tracing the evolutionary histories of ultra-rare variants using variational dating of large ancestral recombination graphs

Ultra-rare variants dominate whole-genome sequencing datasets, yet their interpretation is limited by allele frequency, which provides little information at very low counts and is highly sensitive to uneven ancestry representation. Allele age offers an ancestry-agnostic alternative but existing methods do not scale to biobank-sized cohorts. Here we present a scalable variational algorithm for dating Ancestral Recombination Graphs (ARGs), implemented in tsdate, together with new distributed methods enabling practical biobank-scale ARG inference using tsinfer. Applied to 47,535 genomes from the Genomics England 100,000 Genomes Project, we infer contiguous ARGs spanning 206 Mb and estimate ages for 23.2 million variants, including 11.8 million singletons. ARG-based allele ages remain accurate under extreme sampling imbalance and, in real data, reveal signatures of purifying selection and clinically relevant heterogeneity among variants with identical observed frequencies. Estimates for recent mutations are precise only at large sample sizes, highlighting the information accessible in the haplotype structure of large datasets. Biobank-scale ARGs therefore enable robust, ancestry-agnostic age estimation for ultra-rare variation with broad utility for statistical and clinical genomics.

genetics↗

bioRxiv2025

Mechanistic and evolutionary insights into a family of aminoacyl-tRNA deacylases that protects against canavanine toxicity

Aminoacyl-tRNA deacylases safeguard the accurate translation of the genetic code by hydrolyzing incorrectly synthesized aminoacyl-tRNAs. Canavanyl-tRNA deacylase (CtdA) was recently shown to protect cells against the toxicity of the non-proteinogenic amino acid canavanine, which is synthesized and accumulated by various plants. In most organisms, canavanine is ligated to tRNAArg, causing translation of arginine codons with canavanine. CtdA prevents canavanine toxicity by hydrolyzing canavanyl-tRNAArg. Here, we investigated the function, structure, substrate specificity, phylogenetic distribution, and evolution of CtdA. We show that CtdA is essential to prevent canavanine cytotoxicity in Salmonella enterica, and its heterologous expression can also rescue Escherichia coli. By determining the structure of CtdA, we identified its putative binding pocket and residues that modulate enzymatic activity and specificity. We also found that CtdA displays a relaxed specificity for the aminoacyl moiety substrate as it hydrolyzes arginyl-tRNAArg. Finally, we showed that despite their structural homology, CtdA and the aminoacyl-tRNA hydrolytic domain of phenylalanyl-tRNA synthetase are functionally and evolutionarily divergent. Collectively, these results substantially expand our understanding of the CtdA family, providing new insights into its structure, function, and evolution. Moreover, this work highlights the diverse mechanisms unique to each organism to ensure faithful translation of the genetic code.

biochemistry↗

bioRxiv2025

Tsbrowse: an interactive browser for Ancestral Recombination Graphs

Ancestral Recombination Graphs (ARGs) represent the interwoven paths of genetic ancestry for a set of recombining sequences. The ability to capture the evolutionary history of samples makes ARGs valuable in a wide range of applications in population and statistical genetics. ARG-based approaches are increasingly becoming a part of genetic data analysis pipelines due to breakthroughs enabling ARG inference at biobank-scale. However, there is a lack of visualisation tools, which are crucial for validating inferences and generating hypotheses. We present tsbrowse, an open-source Python web-app for the interactive visualisation of the fundamental building-blocks of ARGs, i.e., nodes, edges and mutations. We demonstrate the application of tsbrowse to various data sources and scenarios, and highlight its key features of browsability along the genome, user interactivity, and scalability to very large sample sizes. AvailabilityPython package: https://pypi.org/project/tsbrowse/, Development version: https://github.com/tskit.dev/tsbrowse, Documentation: https://tskit.dev/tsbrowse/docs/

genomics↗

bioRxiv2023

Jumbo phages are active against extensively-drug-resistant eyedrop-associated Pseudomonas aeruginosa infections.

Antibiotic resistant bacteria present an emerging challenge to human health as the pressure instituted on the microbial world through the liberal use of antibiotics has resulted in their emergence across the globe. Those bacteria that acquire mobile genetic elements such as plasmids are especially concerning because those plasmids may be shared readily with other microbes that then can also become antibiotic resistant. Serious infections have recently been related to contamination of preservative-free eyedrops with extensively drug resistant (XDR) isolates of Pseudomonas aeruginosa, already resulting in three deaths. These drug-resistant isolates cannot be managed with most conventional antibiotics. We sought to identify alternatives to conventional antibiotics for lysis of these XDR isolates, and identified multiple bacteriophages (viruses that attack bacteria) that killed them efficiently. We found both jumbo phages (>200kb in genome size) and non-jumbo phages that were active against these isolates, the former killing more efficiently. Jumbo phages effectively killed the 3 separate XDR P. aeruginosa isolates both on solid and liquid medium. Given the ongoing nature of the XDR P. aeruginosa eyedrop outbreak, the identification of phages active against them provides physicians with several novel potential alternatives for treatment.

microbiology↗

bioRxiv2023

Dual functioning by the PhoR sensor is a key determinant to Mycobacterium tuberculosis virulence

PhoP-PhoR empowers M. tuberculosis to adapt to diverse environmental conditions, and remains essential for virulence. Although PhoP and PhoR have been structurally characterized, the signal(s) that this TCS responds to remains unknown. In this study, we show that PhoR is a sensor of acidic pH/high salt conditions, which activate PhoP via phosphorylation. Transcriptomic studies uncover that acidic pH-inducible expression of PhoP regulon is significantly inhibited in a PhoR-deleted M. tuberculosis. Using genome-wide screening we further identify a non-canonical mechanism of PhoP phosphorylation by the sensor kinase PrrB. To investigate how phosphorylation of PhoP is regulated, we discovered that PhoR functions as a phosphatase. Our results identify the motif/residues responsible for contrasting kinase/phosphatase dual functioning of PhoP, and collectively determine the homeostatic regulation of intra-mycobacterial P~PhoP which controls the final output of PhoP regulon. Together, these data uncover that PhoR plays a central role in mycobacterial adaptation to low pH conditions within the host macrophage phagosome. Consistent with these results a PhoR-deleted M. tuberculosis remains significantly attenuated in macrophages and animal models.

microbiology↗

bioRxiv2022

Bacterial metatranscriptomes in wastewater can differentiate virally infected human populations

Monitoring wastewater samples at building-level resolution screens large populations for SARS-CoV-2, prioritizing testing and isolation efforts. Here we perform untargeted metatranscriptomics on virally-enriched wastewater samples from 10 locations on the UC San Diego campus, demonstrating that resulting bacterial taxonomic and functional profiles discriminate SARS-CoV-2 status even without direct detection of viral transcripts. Our proof-of-principle reveals emergent threats through changes in the human microbiome, suggesting new approaches for untargeted wastewater-based epidemiology.

microbiology↗

bioRxiv2021

Multi-omics profiling of Earth's biomes reveals that microbial and metabolite composition are shaped by the environment

As our understanding of the structure and diversity of the microbial world grows, interpreting its function is of critical interest for understanding and managing the many systems microbes influence. Despite advances in sequencing, lack of standardization challenges comparisons among studies that could provide insight into the structure and function of microbial communities across multiple habitats on a planetary scale. Technical variation among distinct studies without proper standardization of approaches prevents robust meta-analysis. Here, we present a multi-omics, meta-analysis of a novel, diverse set of microbial community samples collected for the Earth Microbiome Project. We include amplicon (16S, 18S, ITS) and shotgun metagenomic sequence data, and untargeted metabolomics data (liquid chromatography-tandem mass spectrometry and gas chromatography mass spectrometry), centering our description on relationships and co-occurrences of microbially-related metabolites and microbial taxa across environments. Standardized protocols and analytical methods for characterizing microbial communities, including assessment of molecular diversity using untargeted metabolomics, facilitate identification of shared microbial and metabolite features, permitting us to explore diversity at extraordinary scale. In addition to a reference database for metagenomic and metabolomic data, we provide a framework for incorporating additional studies, enabling the expansion of existing knowledge in the form of a community resource that will become more valuable with time. To provide examples of applying this database, we outline important ecological questions that can be addressed, and test the hypotheses that every microbe and metabolite is everywhere, but the environment selects. Our results show that metabolite diversity exhibits turnover and nestedness related to both microbial communities and the environment. The relative abundances of microbially-related metabolites vary and co-occur with specific microbial consortia in a habitat-specific manner, and highlight the power of certain chemistry - in particular terpenoids - in distinguishing Earths environments.

ecology↗

bioRxiv2020

Functional assessment of the "two-hit" model for neurodevelopmental defects in Drosophila and X. laevis

We previously identified a deletion on chromosome 16p12.1 that is mostly inherited and associated with multiple neurodevelopmental outcomes, where severely affected probands carried an excess of rare pathogenic variants compared to mildly affected carrier parents. We hypothesized that the 16p12.1 deletion sensitizes the genome for disease, while "second-hits" in the genetic background modulate the phenotypic trajectory. To test this model, we examined how neurodevelopmental defects conferred by knockdown of individual 16p12.1 homologs are modulated by simultaneous knockdown of homologs of "second-hit" genes in Drosophila melanogaster and Xenopus laevis. We observed that knockdown of 16p12.1 homologs affect multiple phenotypic domains, leading to delayed developmental timing, seizure susceptibility, brain alterations, abnormal dendrite and axonal morphology, and cellular proliferation defects. Compared to genes within the 16p11.2 deletion, which has higher de novo occurrence, 16p12.1 homologs were less likely to interact with each other in Drosophila models or a human brain-specific interaction network, suggesting that interactions with "second-hit" genes may confer higher impact towards neurodevelopmental phenotypes. Assessment of 212 pairwise interactions in Drosophila between 16p12.1 homologs and 76 homologs of patient-specific "second-hit" genes (such as ARID1B and CACNA1A), genes within neurodevelopmental pathways (such as PTEN and UBE3A), and transcriptomic targets (such as DSCAM and TRRAP) identified genetic interactions in 63% of the tested pairs. In 11 out of 15 families, homologs of patient-specific "second-hits" enhanced or suppressed the phenotypic effects of one or many 16p12.1 homologs. In fact, homologs of SETD5 synergistically interacted with homologs of MOSMO in both Drosophila and X. laevis, leading to modified cellular and brain phenotypes, as well as axon outgrowth defects that were not observed with knockdown of either individual homolog. Our results suggest that several 16p12.1 genes sensitize the genome towards neurodevelopmental defects, and complex interactions with "second-hit" genes determine the ultimate phenotypic manifestation. Author SummaryCopy-number variants, or deletions and duplications in the genome, are associated with multiple neurodevelopmental disorders. The developmental delay-associated 16p12.1 deletion is mostly inherited, and severely affected children carry an excess of "second-hits" variants compared to mildly affected carrier parents, suggesting that additional variants modulate the clinical manifestation. We studied this "two-hit" model using Drosophila and Xenopus laevis, and systematically tested how homologs of "second-hit" genes modulate neurodevelopmental defects observed for 16p12.1 homologs. We observed that 16p12.1 homologs independently led to multiple neurodevelopmental features and weakly interacted with each other, suggesting that interactions with "second-hit" homologs potentially have a higher impact towards neurodevelopmental defects than interactions between 16p12.1 homologs. We tested 212 pairwise interactions of 16p12.1 homologs with "second-hit" homologs and genes within conserved neurodevelopmental pathways, and observed modulation of neurodevelopmental defects caused by 16p12.1 homologs in 11 out of 15 families, and 16/32 of these changes could be attributed to genetic interactions. Interestingly, we observed that SETD5 homologs interacted with homologs of MOSMO, which conferred additional neuronal phenotypes not observed with knockdown of individual homologs. We propose that the 16p12.1 deletion sensitizes the genome to multiple neurodevelopmental defects, and complex interactions with "second-hit" genes determine the clinical trajectory of the disorder.

genomics↗

bioRxiv2020

Metagenomic characterization of soil microbial communities in the Luquillo experimental forest (Puerto Rico) and implications for nitrogen cycling

The phylogenetic and functional diversity of microbial communities in tropical rainforests, and how these differ from temperate communities remain poorly described but are directly related to the increased fluxes of greenhouse gases such as nitrous oxide (N2O) from the tropics. Towards closing these knowledge gaps, we analyzed replicated shotgun metagenomes representing distinct life zones from four locations in the Luquillo Experimental Forest (LEF), Puerto Rico. These soils had a distinct microbial community composition and lower species diversity when compared to temperate grasslands or agricultural soils. Unlike temperate soils, LEF soils showed little stratification with depth in the first 0-30cm, with ~45% of community composition differences explained solely by location. The relative abundances and nucleotide sequences of N2O reductases (nosZ) were highly similar between tropical forest and temperate soils. However, respiratory NO reductase (norB) was 2-fold more abundant in the tropical soils, which might be relatable to their greater N2O emissions. Nitrogen fixation (nifH) also showed higher relative abundance in rainforest compared to temperate soils (20% vs. 0.1-0.3% of bacterial genomes in each soil type harbored the gene, respectively). Collectively, these results advance our understanding of spatial diversity and metabolic repertoire of tropical rainforest soil communities, and should facilitate future ecological modeling efforts. ImportanceTropical rainforests are the largest terrestrial sinks of atmospheric CO2 and the largest natural source of N2O emissions, two critical greenhouse gases for the climate. The microbial communities of rainforest soils that directly or indirectly, through affecting plant growth, contribute to these fluxes remain poorly described by cultured-independent methods. To close this knowledge gap, the present study applied shotgun metagenomics to samples selected from 3 distinct life zones within the Puerto Rico rainforest. The results advance our understanding of microbial community diversity in rainforest soils and should facilitate future studies of natural or manipulated perturbations of these critical ecosystems.

microbiology↗

bioRxiv2019

Drosophila models of pathogenic copy-number variant genes show global and non-neuronal defects during development

While rare pathogenic copy-number variants (CNVs) are associated with both neuronal and non-neuronal phenotypes, functional studies evaluating these regions have focused on the molecular basis of neuronal defects. We report a systematic functional analysis of non-neuronal defects for homologs of 59 genes within ten pathogenic CNVs and 20 neurodevelopmental genes in Drosophila melanogaster. Using wing-specific knockdown of 136 RNA interference lines, we identified qualitative and quantitative phenotypes in 72/79 homologs, including 21 lines with severe wing defects and six lines with lethality. In fact, we found that 10/31 homologs of CNV genes also showed complete or partial lethality at larval or pupal stages with ubiquitous knockdown. Comparisons between eye and wing-specific knockdown of 37/45 homologs showed both neuronal and non-neuronal defects, but with no correlation in the severity of defects. We further observed disruptions in cell proliferation and apoptosis in larval wing discs for 23/27 homologs, and altered Wnt, Hedgehog and Notch signaling for 9/14 homologs, including AATF/Aatf, PPP4C/Pp4-19C, and KIF11/Klp61F. These findings were further supported by tissue-specific differences in expression patterns of human CNV genes, as well as connectivity of CNV genes to signaling pathway genes in brain, heart and kidney-specific networks. Our findings suggest that multiple genes within each CNV differentially affect both global and tissue-specific developmental processes within conserved pathways, and that their roles are not restricted to neuronal functions. AUTHOR SUMMARYRare copy-number variants (CNVs), or large deletions and duplications in the genome, are associated with both neuronal and non-neuronal clinical features. Previous functional studies for these disorders have primarily focused on understanding the cellular mechanisms for neurological and behavioral phenotypes. To understand how genes within these CNVs contribute to developmental defects in non-neuronal tissues, we assessed 79 homologs of CNV and known neurodevelopmental genes in Drosophila models. We found that most homologs showed developmental defects when knocked down in the adult fly wing, ranging from mild size changes to severe wrinkled wings or lethality. Although a majority of homologs tested showed defects when knocked down specifically in wings and eyes, we found no correlation in the severity of the observed defects in these two tissues. A subset of the homologs showed disruptions in cellular processes in the developing fly wing, including alterations in cell proliferation, apoptosis, and cellular signaling pathways. Furthermore, human CNV genes also showed differences in gene expression patterns and interactions with signaling pathway genes across multiple human tissues. Our findings suggest that genes within CNV disorders affect global developmental processes in both neuronal and non-neuronal tissues.

genomics↗

bioRxiv2019

Genome Repository of Oiled Systems (GROS): an interactive and searchable database that expands the catalogued diversity of crude oil-associated microbes

Indigenous microbial communities ultimately control the fate of petroleum hydrocarbons (PHCs) that enters the natural environment through natural seeps or accidental oil spills, but the interactions among microbes and with their chemical environment during oil biodegradation are highly complex and poorly understood. Genome-resolved metagenomics have the potential to help in unraveling these complex interactions. However, the lack of a comprehensive database that integrates existing genomic/metagenomic data from oiled environments with physicochemical parameters known to regulate the fate of PHCs currently limits data analysis and interpretations. Here, we present a curated, comprehensive, and searchable database that documents microbial populations in oiled ecosystems on a global scale, along with underlying physicochemical data, geocoded via GIS to reveal geographic distribution patterns of the populations. Analysis of the ~2,000 metagenome-assembled genomes (MAGs) available in the database revealed strong ecological niche specialization within habitats e.g., specialization to coastal sediments vs. water-column vs. deep-sea sediments. Over 95% of the recovered MAGs represented novel and uncultured species underscoring the limited representation of cultured organisms from oil-contaminated and oil reservoir ecosystems. The majority of MAGs linked to oiled ecosystems are members of the rare biosphere in non-oiled samples, except for the Gulf of Mexico (GoM) which appears to be primed for oil biodegradation. GROS should facilitate future work toward a more predictive understanding of the microbial taxa and their activities that control the fate of oil spills as well as serve as a model approach for building similar resources for additional environmental processes and omic data of interest.

genomics↗

bioRxiv2018

The influence of rare variants in circulating metabolic biomarkers

Circulating metabolite levels are biomarkers for cardiovascular disease (CVD). We tested association between rare sequence variants and 226 serum lipoproteins, lipids and amino acids in 7,142 healthy participants. Gene-based association analyses identified novel gene-trait associations with ACSL1, MYCN, FBXO36 and B4GALNT3 (p<2.5 x 10-6), and confirmed established associations. Regulation of the pyruvate dehydrogenase (PDH) complex was associated for the first time, in gene set analyses, with IDL and LDL parameters, as well as circulating cholesterol (pMETASKAT <2.41 x 10-6). Individuals at the lower tails of the distributions of four out of 49 lipoproteins and lipids had an excess of predicted deleterious variants in lipoprotein disorder and metabolism gene sets (ppermutation<0.00037). These four traits were CVD risk factors (e.g. S-VLDL-C), demonstrating that rare \"protective\" variation is a significant contributor to lipoprotein levels in a healthy population. In conclusion, rare variant analysis of these important metabolic biomarkers reveals novel loci and pathways involved in their regulation.

genetics↗