Biology subjects

Bawa, P. S.

Publications and source records attributed to Bawa, P. S..

8 recordsLinked to original sources

bioRxiv2026

A non-enzymatic role for METTL3 as an Androgen Receptor co-regulator that promotes prostate cancer proliferation.

Metastatic prostate cancer (PCa) continues to be a major cause of death in males, despite advances in treatment. Most treatment focuses on targeting the Androgen Receptor (AR), the main oncogene responsible for driving most prostate tumors. Despite these therapies targeting AR, the majority of patients still succumb to AR-driven disease. Therefore, there is a critical need for understanding how AR functions to promote prostate cancer growth and identify alternative therapeutic targets in AR-driven PCa. One avenue garnering attention is targeting epigenetic regulators that promote AR-activity; however, the importance of epitranscriptomic regulators, like those that modify mRNAs, is not well understood. Here, we identify a new role for the key catalytic subunit of the RNA N6-methyladenosine (m6A) transferase complex, METTL3, as an AR-coregulator. METTL3 is overexpressed in prostate tumors compared to normal tissue, and METTL3 protein is elevated in AR-expressing cell lines. Depletion of METTL3 significantly reduces proliferation of cancer cells and has no effect on the growth of non-transformed prostate epithelial cells, despite decreasing global m6A levels on mRNA. The catalytic activity of METTL3 is dispensable for the growth of both non-transformed and PCa cell lines, as pharmacologic inhibition of METTL3 does not inhibit proliferation, despite the reduction of global m6A on mRNA. Overexpression of both wild-type and catalytically inactive METTL3 mutants enhances cell viability and rescues cells in which METTL3 is knocked down. Finally, we report on direct interaction between AR and METTL3, their co-localization on chromatin, and reduced AR-cistromic occupancy within cells with METTL3 knockdown. Together, these findings identify a non-enzymatic role for METTL3 in supporting AR-driven transcriptional programs and PCa proliferation.

cancer biology↗

bioRxiv2026

A chimeric human-mouse lung vascular model using induced pluripotent stem cells reveals insights into the pathogenesis of BMPR2-related pulmonary hypertension

Advances in tissue biology have revealed remarkable transcriptomic heterogeneity of endothelial cells between and within organ systems. This necessitates more precise models of organ-specific endothelium to understand the pathogenesis of genetic vascular disorders, such as pulmonary hypertension (PH), where gene-disease associations have implicated endothelial cell dysfunction as a key driver of disease pathogenesis. Towards this end, human induced pluripotent stem cells (hiPSCs) hold immense promise for PH disease modeling where hiPSCs are generated from an affected individual and undergo gene correction to generate syngeneic controls that can be differentiated to endothelial cells (hiEndos), providing a limitless source of material for downstream studies; however, the ability to generate lung-specific hiEndos to model pulmonary vascular disease has been limited. To overcome this challenge, we developed a chimeric human-mouse lung vascular model wherein hiEndos are first patterned via BMP9-induced signaling towards a lung-like molecular phenotype in vitro and are then intravenously transplanted into the mouse lung vasculature in vivo to generate orthotopic lung-specific endothelium for downstream studies. Transplanted pre-patterned hiEndos form functional connections to the native mouse lung vasculature and upregulate differentiated lung-specific molecular cell subtype profiles that include capillary- and arterial-like cell populations. To apply this approach for disease modeling, we generated new hiPSC lines by reprogramming fibroblasts from individuals of the 2001 landmark cohort of BMPR2 gene variant-associated PH and developed a novel in vivo competitive lung endothelial reconstitution assay to quantify functional and molecular differences between human BMPR2-variant vs syngeneic gene-corrected/edited hiEndos. Our approach revealed novel insights into PH disease pathogenesis, not previously evident with prior models, including BMPR2 variant-induced in vivo defects in human lung capillary gene expression, elevated lncRNA H19 expression, increased AHR signaling, and diminished functional capacity to repopulate the pulmonary vascular endothelium.

genetics↗

bioRxiv2026

AT2-intrinsic Z-AAT expression drives conserved inflammatory and proteotoxic stress responses and predisposes to emphysema

Individuals homozygous for the SERPINA1 "Z" mutation with alpha-1 antitrypsin deficiency (AATD) are highly susceptible to emphysema. This predisposition has classically been attributed to a relative deficiency of circulating alpha-1 antitrypsin (AAT) reaching the lungs and associated protease-antiprotease imbalance. Accumulating evidence suggests that the presence of misfolded Z-AAT protein either in the circulation, the lung interstitium, or within resident lung cells could contribute to emphysema pathogenesis. We have shown that type 2 alveolar epithelial cells (AT2s), progenitor cells of the lung alveolus, heterogeneously retain Z-AAT and exhibit a transcriptional disease signature in AATD patient samples. However, a lack of model systems that faithfully recapitulate AT2 biology and associated Z-AAT expression has limited our ability to study this phenomenon. Here, we apply syngeneic induced pluripotent stem cell-derived AT2s (iAT2s) and a novel mouse model featuring AT2-specific inducible human SERPINA1 expression to interrogate the cell-instrinsic consequences of Z-AAT expression, validating findings in an independent dataset of human COPD lung tissue comparing ZZ to MM SERPINA1 genotypes. We find further evidence of Z-AAT retention within AT2s and identify shared AT2 transcriptomic disease signatures conserved across model systems, characterized by innate immune and inflammatory signaling, NF-{kappa}B activation, and endoplasmic reticulum stress. Mice with AT2-specific Z-AAT expression additionally demonstrate increased susceptibility to elastase-induced emphysema, providing functional evidence for AT2-intrinsic contributions to AATD-associated lung disease. Within iAT2s, a subpopulation of Z-AAT expressing cells exhibits activation of the PERK-eIF2 signaling axis and markers of an alveolar basal intermediate (ABI) state, emerging cell-autonomously in the absence of mesenchymal co-culture.Together, these data provide evidence that Z-AAT expression in AT2s induces heterogenous cell-intrinsic stress responses including proteotoxic stress, inflammatory signaling, and aberrant cell fate adoption, and is sufficient to result in predisposition to injury, supporting a potential contribution of AT2-intrinsic Z-AAT toxicity to human AATD-associated emphysema pathogenesis.

cell biology↗

bioRxiv2026

The chromatin remodeling complex PRC2 safeguards cell fate in alveolar epithelial type 2 cells

Maintenance of the gas exchange surface throughout life and regeneration of the lung after injury requires tight regulation of epithelial cell fate and function. Alveolar epithelial type 2 (AT2) cells serve as the progenitors of the distal epithelium, differentiating into alveolar epithelial type 1 (AT1) cells or proliferating to maintain the quorum of AT2 cells. Here we describe the role of the chromatin regulator polycomb repressive complex 2 (PRC2) in the maintenance of AT2 cell fate in the adult alveolus. Cross-species single-cell transcriptomic analyses identified PRC2 activation in proliferative AT2 populations. PRC2 loss of function in human iPSC-derived AT2 (iAT2) cells and primary murine AT2 cells in vitro resulted in loss of AT2 cell state and emergence of programs reminiscent of alveolar-basal intermediate (ABI) cell states, while overexpression of the PRC2 enzymatic component EZH2 in human iAT2 cells augmented the AT2 cell program. Genetic loss of PRC2 function in the AT2 lineage in adult mice in vivo led to emphysematous remodeling of the lung and induced a time-dependent series of transitions of AT2 cells through an alveolar-basal intermediate (ABI) state into Krt5+ basal-like cells. Comparison of murine ABI cells to human disease-associated ABI cells demonstrates de-repression of canonical PRC2 targets during transition to ABI and basal-like states in human fibrosis, implicating PRC2 is a conserved regulator of AT2 cell fate. Together, these findings define PRC2 complex function during AT2 cell self-renewal as a critical guardrail for maintaining epithelial cell fate in the adult lung.

genetics↗

bioRxiv2026

Bidirectional fibrogenic cross-talk revealed in a human iPSC-derived epithelial-mesenchymal co-culture model of pulmonary fibrosis

Pulmonary fibrosis (PF) can arise from mutations in alveolar epithelial type 2 (AT2) cell-specific genes, but manifests in fibrotic activation of mesenchymal cells, thus involving fibrogenic epithelial-mesenchymal crosstalk. The ligand-receptor interactions underlying the onset and early progression of PF remain poorly understood. Induced pluripotent stem cell (iPSC)-derived models are powerful tools to study respiratory diseases, yet are currently limited to reductionist single lineage epithelial models or multi-lineage systems that lack purity and lung-specificity of the mesenchyme. Here we generate a human iPSC line carrying both a lung mesenchyme-specific reporter (TBX4-LERtdTomato) and a reporter for mesenchymal activation/differentiation (ACTA2GFP). Applying this line, we develop a directed differentiation protocol capable of generating cells that express key molecular and functional features of primary human developing lung mesenchyme across multiple iPSC genetic backgrounds. We then establish co-cultures of these iPSC-derived lung mesenchymal cells (iLM) with patient-specific iPSC-derived alveolar epithelial type 2 cells (iAT2s) carrying an SFTPCI73T mutation as a model for PF. We find increased expression of fibrotic markers in co-cultures with mutant iAT2s as compared to co-cultures with gene-corrected iAT2s. Moreover, mutant iAT2s express markers of alveolar-basal intermediate (ABI) cells only in the presence of iLM, suggesting that bidirectional crosstalk promotes this aberrant cell state. We identify ligand-receptor pairs enriched in co-cultures with mutant iAT2s, including TGF{beta}, multiple integrins, and additional genes that have not been previously linked to PF. Finally, we show that small molecule-mediated inhibition of TGF{beta} or integrins V{beta}1/V{beta}6 attenuates both fibrotic mesenchymal activation and the presence of ABI cells in iLM/iAT2 co-cultures. Thus, we have established a human iPSC-derived co-culture system that recapitulates key molecular hallmarks of bidirectional fibrogenic epithelial-mesenchymal crosstalk in pulmonary fibrosis, and enables the identification and study of potentially druggable pathways involved in disease initiation and progression.

molecular biology↗

bioRxiv2025

Generation of effector CD4+ T cells from Human iPSC

Off the shelf CD4+ T cell therapies, particularly those with immunoregulatory or cell repair functions, could be transformative in CAR therapies for cancers and treatment of chronic inflammatory diseases. However, progress is stunted in this area due to challenges generating human CD4+ T cells from induced pluripotent stem cells. Here we describe a key role for the withdrawal of Notch ligand during the final step of stimulation through the T cell receptor to prompt T cell maturation allowing access to the CD4 lineage in iPSC T cells (iCD4+ T cells). Functional analyses of iCD4+ T cells using a novel high-parameter CyTOF intracellular cytokine staining panel revealed both canonical Th1 cytokine signatures and cells producing varying combinations of other cytokines including IL-4, IL-8, and IL-13. Single cell RNA sequencing of iCD4+ T cells demonstrated a transcriptional signature similar to human blood CD4+ T cells. We believe this robust yet simple platform represents a key step towards the generation of off the shelf iCD4+ T cell therapies with utility for the treatment of a panoply of diseases including cancer and inflammatory autoimmune disorders. HIGHLIGHTST cell receptor stimulation of iPSC CD4+/CD8+ T cell progenitors on coating without Notch ligand allows access to the CD4+ T cell lineage iPSC derived CD4+ T cells express varied cytokines in response to stimulation

immunology↗

bioRxiv2025

A stem cell-based platform for functional analysis of genetic variants in lung disease

Advances in genetic and transcriptomic technologies have identified large numbers of genes and variants of potential importance to human disease. Determining the function of these genes and variants is a critical bottleneck in understanding disease etiology. Variants of uncertain significance (VUS) are highly prevalent in our genomes, but our ability to identify them significantly outpaces our ability to determine their molecular and clinical consequences. We developed a genetically tractable induced pluripotent stem cell (iPSC) based platform to investigate gene variant pathogenicity in lung disease, using primary ciliary dyskinesia (PCD) as a model. We identified an individual with a clinical diagnosis of PCD and a VUS in the gene Multiciliate differentiation and DNA synthesis associated cell cycle protein (MCIDAS). Through gene-editing of iPSC-derived airway basal stem cells (iBCs), we precisely defined the molecular and cellular pathogenicity of the variant providing a successful application of the iPSC system to diagnose a lung disease.

cell biology↗

bioRxiv2025

SOX2 utilizes FOXA1 as a heteromeric transcriptional partner to drive proliferation in therapy-resistant prostate cancer

Treatment options and diagnostic outlook for men with advanced, therapy resistant prostate cancer (PCa) are extremely poor; this is primarily due to the common lack of durable response to androgen receptor (AR) targeted therapies and phenotypic transdifferentiation into a particularly lethal subtype known as neuroendocrine prostate cancer (NEPC). In this study, we mechanistically determine that SOX2 (a transcription factor originally repressed by AR) physically binds and acts in a concerted manner with FOXA1 (a key AR pioneering cofactor) to regulate a subset of genes which promote cell cycle progression, and lineage plasticity in AR-refractory prostate cancers. Our findings assert the SOX2/FOXA1 interaction as an important mediator of resistance to AR-targeted therapy and a driver of NEPC and lineage plasticity; their coordinated action and downstream signaling offers a potential novel therapeutic opportunity in late-stage PCa.

cancer biology↗