Biology subjects

Chandra, D.

Publications and source records attributed to Chandra, D..

7 recordsLinked to original sources

bioRxiv2026

Epithelial QKI Protects Against Emphysema by Maintaining Mitochondrial Integrity

Single-cell transcriptomic profiling of chronic obstructive pulmonary disease (COPD) lungs identified QKI, an RNA-binding protein, as a candidate emphysema-associated gene, but its epithelial role in COPD pathobiology remains unclear. We show that QKI expression is reduced in human COPD lungs and that alveolar type 2 epithelial (AT2) cell QKI protein levels correlate strongly with spirometric indices and diffusing capacity (DLCO). Lung epithelium-specific QKI knockout mice (QKI{Delta}/{Delta}) developed spontaneous airspace enlargement with emphysema-like mechanics, and QKI-deficient AT2 cells showed impaired spheroid colony formation and increased apoptosis. Integrated transcriptomic and proteomic analyses of primary AT2 cells revealed a selective reduction in functional mitochondrial (respiratory-chain and metabolic) protein abundance despite relatively preserved transcript levels, consistent with mitochondrial transcriptome-proteome discordance. QKI loss increased mtDNA abundance and TOMM20 staining but decreased ATP5A, indicating accumulation of structurally increased but functionally dysfunctional mitochondria. In human epithelial cells, CRISPR-mediated QKI deficiency reduced oxidative respiration, increased glycolytic reliance, elevated mitochondrial ROS and membrane potential, and increased apoptosis; these phenotypes were partially rescued by QKI re-expression. These findings identify epithelial QKI as a regulator of mitochondrial integrity and stress tolerance in COPD.

cell biology↗

bioRxiv2026

Coregulation by arcA and fnr protects Salmonella Typhimurium from bile stress by maintaining redox homeostasis and membrane integrity

Bile salts constitute a major antimicrobial barrier encountered by Salmonella Typhimurium during infections. Here, we identified the interplay of two global regulators, Fnr (fumarate and nitrate reductase) and ArcA (aerobic respiration control protein A), in determining adaptive responses to bile. We utilized WT, {Delta}fnr, {Delta}arcA and {Delta}arcA{Delta}fnr strains and compared the expression and functional responses of S. Typhimurium to bile. The highlights of this study are: First, the {Delta}arcA and {Delta}arcA{Delta}fnr strain form smaller colonies on LB agar plates and the {Delta}arcA{Delta}fnr strain displays lower motility. Second, qRT-PCR expression analysis demonstrates that fnr is induced early with bile, followed by arcA. Also, arcA transcripts are lower in the {Delta}fnr strain and fnr expression is also partially lower in the {Delta}arcA strain. Third, the {Delta}arcA and {Delta}fnr strains display partial sensitivity to bile, whereas the {Delta}arcA{Delta}fnr strain exhibits hypersensitivity to bile. Upon bile exposure, the {Delta}arcA{Delta}fnr strain displays elevated transcripts of the major antioxidant genes (sodA and katG) and outer membrane protein (ompC), higher induction of reactive oxygen species (ROS), and greater membrane damage. Fourth, intracellular nitrite is induced earlier than ROS with bile. Fifth, arcA and fnr protects S. Typhimurium from bile induced stress by activating the nitrate metabolism pathway, which lowers ROS. Functionally, pretreatment of the deletion strains with sodium nitrate reduces ROS, improves membrane integrity and survival with bile. Overall, these findings demonstrate that arcA and fnr function cooperatively to utilize alternate electron acceptors, reduce dependence on aerobic respiration and lower ROS to improve survival of S. Typhimurium during bile stress.

microbiology↗

bioRxiv2025

Suppressing microtubule detyrosination augments AAV2 endosomal escape and gene delivery.

Adeno-associated virus (AAV) is a widely used vector for gene delivery, yet the host intracellular trafficking barriers often limit its efficacy. Here, we identify microtubule detyrosination--a tubulin post-translational modification--as a key regulator of AAV2 endo-lysosomal processing. Using super-resolution microscopy (SIM/STORM), we show that upon AAV2 endocytosis, the host upregulates microtubule detyrosination via GSK3{beta}-CLASP2 signaling axis. Single particle tracking of the virus reveals that detyrosinated microtubules form a physical and functional barrier, restricting AAV2 motility and promoting lysosomal trapping. Restoring microtubule tyrosination--via tubulin-tyrosine ligase overexpression or pharmacological inhibition of detyrosination with parthenolide--boosted AAV2 endosomal escape, perinuclear accumulation, and gene delivery in cells. Notably, a clinically relevant prodrug of parthenolide, DMAPT, also displayed a similar trend of enhanced AAV2-driven factor IX expression in hemophilia B mouse models. Our findings uncover a host mechanism that reshapes the microtubule landscape to restrict AAV2 trafficking and identify microtubule detyrosination as a novel druggable target to improve AAV2-based gene therapies.

cell biology↗

bioRxiv2025

Reciprocal Regulation Between the SCFFBXO24 Ubiquitin E3 Ligase and FoxP1 Protein

Forkhead Box Protein P1 (FoxP1) is a crucial transcriptional repressor essential for the development of the brain and heart. In adults, FoxP1 protein levels are dysregulated in a variety of disorders, including chronic obstructive pulmonary disease (COPD), atherosclerosis, and heart failure, where they causally contribute to disease pathogenesis. Although independent investigators have reported that FoxP1 protein is ubiquitinated, and E3 ligases have been identified for other FoxP family proteins, the identity of the E3 ligase that controls FoxP1 protein stability has remained unknown. Here, we identify FBXO24, a subunit of the Skp-Cullin-F-box (SCF) ubiquitin E3 ligase complex, as the regulator of FoxP1 ubiquitination and stability. Specifically, FBXO24 regulates K48 and K63 ubiquitination, complexes with, and co-localizes to the nucleus with FoxP1 protein in lung epithelial cells. Depleting FBXO24 reverses the unfolded protein response and cell death triggered by loss of FoxP1 protein in lung epithelium, suggesting a protective role. Additionally, FBXO24 knockout mice exhibit elevated FoxP1 levels in the lung and heart and reduced unfolded protein response activity after short-term cigarette smoke exposure. Intriguingly, we also uncovered bidirectional regulation, whereby FoxP1 protein binds to the FBXO24 promoter to suppress FBXO24 transcription. To our knowledge, this is the first evidence that a substrate for an E3 ligase can also regulate the E3 ligase and, therefore, control levels of other substrates, revealing new regulatory networks. Targeting FBXO24 may offer a therapeutic strategy for COPD, atherosclerosis, and heart failure by stabilizing FoxP1 levels in the heart and lungs and mitigating harmful downstream effects.

molecular biology↗

bioRxiv2023

Global transcriptome analysis reveals Salmonella Typhimurium employs the nitrate-dependent anaerobic pathway to combat bile stress

Salmonella Typhimurium is an enteric pathogen that is highly tolerant to bile. Next-generation mRNA sequencing was performed to analyse the stress and adaptive responses of S. Typhimurium to bile. We identified the cellular pathways affected during bile stress in wild type (WT) and a mutant lacking cspE ({Delta}cspE), which plays an essential role in protection from bile stress. We observed transcriptional upregulation of several genes involved in nitrate metabolism, in response to bile stress. These genes were also differentially expressed between the bile-resilient WT and the bile-sensitive {Delta}cspE strain. To understand the role of nitrate metabolism in bile stress response, we generated a strain lacking fnr ({Delta}fnr), which is the global regulator of nitrate metabolism in S. Typhimurium. fnr was highly induced in the bile treated WT strain but not in the {Delta}cspE strain. Notably, the {Delta}fnr strain was susceptible to bile-mediated killing. Our studies revealed a new role for fnr in mediating the bile stress response. In addition, a strain lacking arcA ({Delta}arcA), a two-component system response regulator involved in anaerobic metabolism, also showed a marked reduction in growth in presence of bile. This corroborated the significance of anaerobic metabolism in S. Typhimurium bile tolerance. Importantly, overexpression of fnr and arcA lowered reactive oxygen species and significantly enhanced the survival of the bile-sensitive {Delta}cspE strain. We also observed that S. Typhimurium pre-treated with nitrate displayed better growth in the presence of bile. Together, these results demonstrate that nitrate-dependent anaerobic metabolism promotes adaptation of S. Typhimurium to bile. ImportanceSalmonella Typhimurium, as an enteric pathogen, manifests an extreme example of bile tolerance. This study describes the diverse metabolic changes at the level of transcriptome in S. Typhimurium exposed to bile. We identified the differential expression of several genes involved in anaerobic metabolism between bile-tolerant WT and bile-sensitive {Delta}cspE strains. Two major regulators of anaerobic metabolism, fnr and arcA, support the growth of S. Typhimurium in bile. Our results highlight that, in presence of bile, S. Typhimurium activates genes involved in anaerobic metabolism, specifically nitrate metabolism, that improves survival of bacteria during bile stress.

microbiology↗

bioRxiv2020

Profiling Active Enzymes for Polysorbate Degradation in Biotherapeutics by Activity-Based Protein Profiling

Polysorbate is widely used to maintain stability of biotherapeutic proteins in formulation development. Degradation of polysorbate can lead to particle formation in drug products, which is a major quality concern and potential patient risk factor. Enzymatic activity from residual enzymes such as lipases and esterases can cause polysorbate degradation. Their high activity, even at low concentration, constitutes a major analytical challenge. In this study, we evaluated and optimized the activity-based protein profiling (ABPP) approach to identify active enzymes responsible for polysorbate degradation. Using chemical probes to enrich active serine hydrolases, more than 80 proteins were identified in harvested cell culture fluid (HCCF) from monoclonal antibodies (mAb) production. A total of 8 known lipases were identified by ABPP, while only 5 lipases were identified by a traditional abundance-based proteomics (TABP) approach. Interestingly, phospholipase B-like 2 (PLBL2), a well-known problematic HCP was not found to be active in process-intermediates from two different mAbs. In a proof-of-concept study, phospholipase A2 group VII (PLA2G7) and sialic acid acetylesterase (SIAE) were identified by ABPP as possible root causes of polysorbate-80 degradation. The established ABBP approach can fill the gap between lipase abundance and activity, which enables more meaningful polysorbate degradation investigations for biotherapeutic development.

bioengineering↗

bioRxiv2019

A new immunotherapeutic concept for pancreatic cancer using childhood vaccine recall antigens and Listeria

Pancreatic ductal adenocarcinoma is highly metastatic, poorly immunogenic, and immune suppression prevents T cell activation in the tumor microenvironment. We developed a microbial-based immunotherapeutic concept for selective delivery of a highly immunogenic tetanus toxoid protein (TT856-1313), into tumor cells by attenuated Listeria monocytogenes, and reactivation of pre-existing TT-specific memory T cells (generated during childhood) to kill infected tumor cells. Thus, TT here functions as an alternative for neoantigens. Treatment of KPC mice with Listeria-TT resulted in TT accumulation in tumors and inside tumor cells, and attraction of predominantly TT-specific memory CD4 T cells. Moreover, gemcitabine (GEM) combined with Listeria-TT significantly improved the migration of CD4 T cells into tumors and the production of perforin and granzyme B, turning cold tumors into immunological hot tumors. In vivo depletion of T cells in Listeria-TT+GEM-treated mice demonstrated CD4 T cell-mediated eradication of tumors and metastases (Mann-Whitney p<0.05). In addition, peritumoral lymph node like structures (LNS) were observed in close contact with the pancreatic tumors displaying CD4 T cells and CD8 T cells of KPC mice treated with Listeria-TT or Listeria-TT+GEM. The production of perforin and granzyme B was observed in LNS of KPC mice that received Listeria-TT+GEM. This combination not only reduced tumor burden (80%) and metastases (87%) significantly (p<0.05, Mann-Whitney), but also improved the survival time of KPC mice with advanced pancreatic cancer substantially (Mantel-Cox p<0.0001). Our results unveil new mechanisms of Listeria and GEM improving immunotherapy for PDAC.

immunology↗