Biology subjects

Rao, S. D.

Publications and source records attributed to Rao, S. D..

5 recordsLinked to original sources

bioRxiv2022

Cytosolic peptide accumulation activates the NLRP1 and CARD8 inflammasomes

NLRP1 and CARD8 are related sensors that form inflammasomes, but the danger signals that they detect are not fully established. These proteins undergo autoproteolysis, generating repressive N-terminal (NT) and inflammatory C-terminal (CT) fragments. The proteasome- mediated degradation of the NT releases the CT from autoinhibition, but the CT is then sequestered in a complex with the full-length sensor and DPP9. Here, we show that cytosolic peptide accumulation activates these inflammasomes. We found that a diverse array of peptides accelerates NT degradation, and those with N-terminal XP sequences also destabilize the ternary complexes. Peptides interfere with many biological processes, including protein folding. We show that unrelated agents that disrupt protein folding also induce NT degradation, but do not cause inflammasome activation because DPP9 sequesters the CT fragments in the absence of XP peptides. Overall, these results indicate that NLRP1 and CARD8 detect protein misfolding that is associated with peptide accumulation.

immunology↗

bioRxiv2021

Oxidized thioredoxin-1 restrains the NLRP1 inflammasome

At least six human proteins detect danger-associated signals, assemble into complexes called inflammasomes, and trigger pyroptotic cell death. NLRP1 was the first protein discovered to form an inflammasome, but the danger signals and molecular mechanisms that control its activation have not yet been fully established. Here, we report that the NACHT-LRR region of NLRP1 directly binds to oxidized form of thioredoxin-1 (TRX1). We found that NLRP1 requires the ATPase activity of its NACHT domain to associate with TRX1, and that this interaction represses inflammasome activation. Moreover, we discovered that several patient-derived missense mutations in the NACHT-LRR region of NLRP1 weaken TRX1 binding, leading to inflammasome hyperactivation and autoinflammatory disease. Overall, our results establish that oxidized TRX1 binds to and restrains the NLRP1 inflammasome, thereby revealing a link between the cellular redox environment and innate immunity.

immunology↗

bioRxiv2021

Independent control of mean and noise by convolution of gene expression distributions

Gene expression noise can reduce cellular fitness or facilitate processes such as alternative metabolism, antibiotic resistance, and differentiation. Unfortunately, efforts to study the impacts of noise have been hampered by a scaling relationship between noise and expression level from a single promoter. Here, we use theory to demonstrate that mean and noise can be controlled independently by expressing two copies of a gene from separate inducible promoters in the same cell. We engineer low and high noise inducible promoters to validate this result in Escherichia coli, and develop a model that predicts the experimental distributions. Finally, we use our method to reveal that the response of a promoter to a repressor is less sensitive with higher repressor noise and explain this result using a law from probability theory. Our approach can be applied to investigate the effects of noise on diverse biological pathways or program cellular heterogeneity for synthetic biology applications.

synthetic biology↗

bioRxiv2020

Chaperone-mediated stress-sensing in Mycobacterium tuberculosis enables fast activation and sustained response

Dynamical properties of gene-regulatory networks are tuned to ensure bacterial survival. In mycobacteria, MprAB-{sigma}E network responds to the presence of stressors, such as surfactants causing surface stress. Positive feedback loops in this network were previously predicted to cause hysteresis, i.e. different responses to identical stressor levels for pre-stressed and unstressed cells. Here we show that hysteresis does not occur in non-pathogenic Mycobacterium smegmatis but occurs in Mycobacterium tuberculosis. However, the observed rapid temporal response in M. tuberculosis is inconsistent with the model predictions. To reconcile these observations, we implement a recently proposed mechanism for stress-sensing: the release of MprB from the inhibitory complex with chaperone DnaK upon the stress exposure. Using modeling and parameter fitting, we demonstrate that this mechanism can accurately describe the experimental observations. Furthermore, we predict perturbations in DnaK expression that can strongly affect dynamical properties. Experiments with these perturbations agree with model predictions, confirming the role of DnaK in fast and sustained response. IMPORTANCEGene-regulatory networks controlling stress response in mycobacterial species have been linked to persistence switches enabling the bacterial dormancy within a host. However, the mechanistic basis of switching and stress sensing is not fully understood. In this paper, combining quantitative experiments and mathematical modeling, we uncover how interactions between two master regulators of stress response -- MprAB two-component system and alternative sigma factor{sigma} E - shape the dynamical properties of surface-stress network. The result show hysteresis (history dependence) in the response of pathogenic bacteria M. tuberculosis to surface stress and lack of the hysteresis in non-pathogenic M. smegmatis. Furthermore, to resolve the apparent contradiction between the existence of hysteresis and fast activation of the response, we utilize a recently proposed role of chaperone DnaK in stress-sensing. The results leads to a novel system-level understanding of bacterial stress-response dynamics.

systems biology↗

bioRxiv2019

Human caspase-1 autoproteolysis is required for ASC-dependent and -independent inflammasome activation

Pathogen-related signals induce a number of cytosolic pattern-recognition receptors (PRRs) to form canonical inflammasomes, which activate pro-caspase-1 and trigger pyroptotic cell death. All well-studied PRRs oligomerize with the pro-caspase-1-adapter protein ASC to generate a single large structure in the cytosol, which induces the autoproteolysis and activation of the pro-caspase-1 zymogen. However, several PRRs can also directly interact with pro-caspase-1 without ASC, forming much smaller \"ASC-independent\" inflammasomes. It is currently thought that pro-caspase-1 autoproteolysis does not occur during, and is not required for, ASC-independent inflammasome activation. Here, we show that the related human PRRs NLRP1 and CARD8 exclusively form ASC-dependent and ASC-independent inflammasomes, respectively, identifying CARD8 as the first PRR that cannot form an ASC-containing signaling platform. Despite their different structures, we discovered that both the NLRP1 and CARD8 inflammasomes require pro-caspase-1 autoproteolysis between the small and large catalytic subunits to induce pyroptosis. Thus, pro-caspase-1 self-cleavage is an obligate regulatory step in the activation of human canonical inflammasomes.

immunology↗