Biology subjects

Rishik, S.

Publications and source records attributed to Rishik, S..

5 recordsLinked to original sources

bioRxiv2026

Sex-divergent responses to microglial depletion suggest distinct regulatory dependencies in the aged brain

Microglia are essential for brain homeostasis, yet their roles in the aged brain remain poorly defined. Using microRNA (miRNA) profiling, cellular-resolution spatial transcriptomics, and bulk proteomics in 21-month-old mice, we characterize sex-dimorphic responses to microglial depletion via CSF1R inhibition (PLX5622 treatment). Microglia-enriched miRNAs, notably miR-146a-5p and miR-223-3p, were downregulated across different brain regions in both sexes. Transcriptional responses were sex dimorphic: females showed predominantly cell-type-specific downregulation, while males showed bidirectional changes including upregulation of Lzts3, Shank3, and Fgfbp1 alongside downregulation of Ang. Proteomic changes were larger in magnitude and independent from mRNA changes: males exhibited 295 differentially expressed proteins (DEPs) versus 34 in females (8.7-fold difference). Male DEPs had opposing directional shifts in synaptic vesicle proteins (upregulated) and mitochondrial ATP synthesis machinery (downregulated). These data describe sex-dimorphic molecular consequences of microglial loss in the aged brain and identify candidate post-transcriptional mechanisms warranting further investigation.

neuroscience↗

bioRxiv2026

Gene amplification during differentiation of mesenchymal stem cells towards chondrocytes

For decades gene amplifications were described as an attribute of tumor cells and as a physiological mechanism to increase gene copy numbers for the higher protein demand during development of amphibians and flies. An increasing number of publications describe gene amplifications in normal mammalian cells during differentiation. Many amplified genes detected in tumor cells overlap with amplified genes detected during stem cell differentiation. Since stem cells have a valuable potency in regenerative therapies and since cartilage regeneration is a highly demanded therapeutic strategy, we investigated gene amplification dynamics during chondrogenic differentiation of human mesenchymal stem cells (hMSCs). Using quantitative PCR, we analyzed copy number changes for genes previously implicated in differentiation as well as genes amplified in chondrosarcoma including CDK4, MDM2, AGAP2, CPT1B, SHANK3, TRIB1, and MYC. Amplifications were transient and stage-specific: CDK4, CPT1B, and SHANK3 exhibited the highest copy number increases at day 2, followed by a gradual decline by day 7, while AGAP2 and MDM2 increased later in differentiation. Laser microdissection of toluidine blue-stained areas revealed heterogeneity in amplification patterns: CDK4 amplification was prominent in areas lacking or showing moderate proteoglycan deposition; CPT1B amplification occurred in regions with absent, moderate, or intense proteoglycan deposition; and SHANK3 amplification was restricted to areas with intense proteoglycan deposition. Notably, regions with the strongest proteoglycan staining exhibited no gene amplification, suggesting that gene amplification is an early, transient event that diminishes as differentiation progresses. These findings highlight gene amplification as a mechanism during chondrogenesis, potentially critical for early differentiation stages and genome stability in mature cells.

Developmental Biology↗

bioRxiv2026

Aging Rhesus Macaque show tissue and sex-specific balance of drifting and coordinated miRNA programs

Macaques research centrality makes it critical to study their molecular aging. We accomplish this for their non-coding transcriptome by sequencing small RNA from 11 organs, with special focus on brain by including 24 brain regions, sampling males and females between ages 3-35 years. Heart, adrenal gland, corpus callosum and caudate putamen showed the most age-deregulated miRNA trajectories. The MIR-154 family, inside the imprinted, rejuvenation-associated Dlk1-Dio3 cluster, was particularly vulnerable. Known age-associated miRNA families LET-7, MIR-29, MIR-17 and MIR-92 were strongly deregulated, with heavy dependence on tissue and sex. MiRNA genomic clusters deregulation was concordant within tissue-sex combinations, implicating upstream regulation rather than random noise. Cross-species comparison with mouse showed ancient miRNAs dominating age-deregulated trajectories. Deregulation direction in tissues-sex was conserved between species at family/cluster levels, but conservation substantially weakened at individual miRNA level. Thus, we mark a decisive step in translating miRNA aging trajectories between two heavily used model organisms. Key FindingsO_LIHeart, adrenal gland, corpus callosum, caudate putamen are hotspots of miRNA age deregulation, with dramatic influence from sex. C_LIO_LINon-brain organs show tissue specific miRNA change, with inconsistent overlap between tissues. C_LIO_LIGenomic clusters of miRNAs were found to be concordant in their age deregulation direction, dependent on tissue and sex, suggesting upstream regulation. C_LIO_LIThe MIR-154 family, housed inside the heavily imprinted Dlk1-Dio3 cluster and processed from the rejuvenation associated MEG3-MIRG host gene is prominently involved in both non-brain organs and brain regions. C_LIO_LIConcentration of age deregulation in evolutionarily ancient miRNAs across species implies regulatory program rather than epigenetic drift, involving MIR-154, LET-7, MIR-29, MIR-17 and MIR-92 families. C_LIO_LIDirection of change conserved between species at the family / genomic cluster level but diminished substantially at individual miRNA level. C_LI

systems biology↗

bioRxiv2026

Physiological re-replication during human stem cell differentiation

During defined developmental windows in Drosophila, controlled re-replication generates physiological gene amplification. Although gene amplification has also been observed during human stem cell differentiation, re-replication in human cells has largely been linked to tumor-associated genome instability. Here, we demonstrate that re-replication likewise operates as a physiological mechanism in human stem cells. Using Rerep-Seq and DNA fiber-combing, we identify distinct phases of re-replication during the differentiation of human myoblasts into myotubes and during the lineage commitment of mesenchymal stem cells toward adipogenic, osteogenic, chondrogenic, and neuronal fates. In all differentiation systems examined, re-replication occurred within defined temporal windows. FACS-isolated re-replicating cells exhibited elevated gene expression using RNA-Seq specifically within re-replicated genomic regions. Moreover, re-replicated DNA was detected as extranuclear DNA. These findings support a model in which cells that do not undergo re-replication, and thus avoid increased chromosomal instability, may nonetheless boost the expression of differentiation-relevant genes by acquiring re-replicated DNA released from neighboring re-replicating cells. We propose that human stem cells exploit an evolutionarily conserved re-replication mechanism to transiently increase gene copy number and thereby meet the heightened protein demands associated with differentiation.

genomics↗

bioRxiv2025

Extracellular vesicles and their RNA cargo facilitate bidirectional cross-kingdom communication between human and bacterial cells

While extracellular vesicles (EVs) are established mediators of intra-species signaling, their role as active participants in cross-kingdom communication remains incompletely understood. Here, we reveal that human colon cells and both Gram-positive and Gram-negative gut bacteria engage in species-specific, EV-mediated molecular dialogue, driven in part by RNA cargo. We show that bacterial EVs (BEVs) induce distinct transcriptomic responses in human cells, and that BEV-RNA independently causes similar effects. Conversely, we demonstrate that human EVs and highly abundant miR-192-5p are differentially internalized by bacteria, affecting their physiology. Our findings support a conceptual model in which EVs function as directional messengers that shape host-microbiome interactions. This study introduces a framework for understanding EVs as cross-kingdom regulators and underscores the importance of tailored, context-specific analyses for understanding the scope of EV-mediated interactions in microbiome-host homeostasis and disease. Highlights[1] L. casei, E. faecalis and P. mirabilis produce BEVs that are internalized by Caco-2 cells at different rates. BEVs produced by L. casei have a positive influence on the viability of Caco-2 cells. Incubation of Caco-2 cells with BEVs leads to changes in the gene expression of immune-response-related genes. [2] BEVs carry RNAs and the type of RNA cargo varies significantly between the BEVs from the different bacteria. Comparison of Caco-2 gene deregulation between BEVs and transfection of RNA isolated from BEV highlights component-specific effects. [3] Caco-2 EVs are taken up by E. faecalis and influence their growth. MiRNA-192-5p can be frequently detected in EVs from Caco-2 cells. Synthetic miR-192-5p is internalized by P. mirabilis and the ability to take up human miRNAs by L. casei and E. faecalis can be increased by packaging of the miRNA in artificial liposomes.

molecular biology↗