Biology subjects

Galapate, C. M.

Publications and source records attributed to Galapate, C. M..

3 recordsLinked to original sources

bioRxiv2023

The lung employs an intrinsic surfactant-mediated inflammatory response for viral defense

Severe Acute Respiratory Syndrome Coronavirus-2 (SARS-CoV-2) causes an acute respiratory distress syndrome (ARDS) that resembles surfactant deficient RDS. Using a novel multi-cell type, human induced pluripotent stem cell (hiPSC)-derived lung organoid (LO) system, validated against primary lung cells, we found that inflammatory cytokine/chemokine production and interferon (IFN) responses are dynamically regulated autonomously within the lung following SARS-CoV-2 infection, an intrinsic defense mechanism mediated by surfactant proteins (SP). Single cell RNA sequencing revealed broad infectability of most lung cell types through canonical (ACE2) and non-canonical (endocytotic) viral entry routes. SARS-CoV-2 triggers rapid apoptosis, impairing viral dissemination. In the absence of surfactant protein B (SP-B), resistance to infection was impaired and cytokine/chemokine production and IFN responses were modulated. Exogenous surfactant, recombinant SP-B, or genomic correction of the SP-B deletion restored resistance to SARS-CoV-2 and improved viability.

immunology↗

bioRxiv2022

Glutamine mimicry suppresses tumor progression through asparagine metabolism in pancreatic ductal adenocarcinoma

In pancreatic ductal adenocarcinoma (PDAC), glutamine is a critical nutrient that drives a wide array of metabolic and biosynthetic processes that support tumor growth. Despite this established dependency, the targeting of specific enzymes involved in glutamine metabolism is yet to yield any clinical benefit. Here, we have examined the therapeutic potential of 6-diazo-5-oxo-L-norleucine (DON), a glutamine antagonist that broadly inhibits glutamine metabolism. We found that DON treatment significantly blocks PDAC tumor growth and attenuates metastasis. Interestingly, we link the effectiveness of DON in PDAC to asparagine (Asn) metabolism. By inhibiting asparagine synthetase (ASNS), DON significantly reduces intracellular Asn production and Asn supplementation rescues the anti-proliferative effects of DON. We discern that PDAC cells upregulate expression of ASNS as a metabolic adaptation and that modulating ASNS levels can impact DON efficacy. Strikingly, in patient-derived organoids, DON responsiveness is inversely correlated with ASNS expression, a feature that is not observed for other metabolic enzymes targeted by DON. We find that treatment with L-asparaginase (ASNase), an enzyme that catabolizes free Asn, synergizes with DON to impact the viability of PDAC cells. Finally, we identify that a combination therapy of DON and ASNase has a significant impact on metastasis. These results shed light on the mechanisms that drive the effects of glutamine mimicry and point to the utility of co-targeting adaptive responses to control PDAC progression.

cancer biology↗

bioRxiv2021

Monitoring Protein Import into the Endoplasmic Reticulum in Living Cells with Proximity Labeling

The proper trafficking of eukaryotic proteins is essential to cellular function. Genetic, environmental, and other stresses can induce protein mistargeting, and in turn threaten cellular protein homeostasis. Current methods for measuring protein mistargeting are difficult to translate to living cells, and thus the role of cellular signaling networks in stress-dependent protein mistargeting processes, such as ER pre-emptive quality control (ER pQC), are difficult to parse. Herein, we use genetically encoded peroxidases to characterize protein import into the endoplasmic reticulum (ER). We show that the ERHRP/cytAPEX pair provides good selectivity and sensitivity for identifying protein mistargeting, using the known ER pQC substrate transthyretin (TTR). Although ERHRP labeling induces formation of detergent-resistant TTR aggregates, this is minimized by using low ERHRP expression, without loss of labeling efficiency. cytAPEX labeling recovers TTR that is mistargeted as a consequence of Sec61 inhibition or ER stress-induced ER pQC. Furthermore, we demonstrate that stress-free activation of the ER stress-associated transcription factor ATF6 recapitulates the TTR import deficiency of ER pQC. Hence, proximity labeling is an effective strategy for characterizing factors that influence ER protein import in living cells.

molecular biology↗