Selectivity among anti-σ factors by Mycobacterium tuberculosis ClpX influences intracellular levels of Extracytoplasmic Function σ factors

Abstract

Extracytoplasmic Function {sigma} factors that are stress inducible are often sequestered in an inactive complex with a membrane-associated anti-{sigma} factor. M. tuberculosis membrane-associated anti-{sigma} factors have a small stable RNA gene A-like degron for targeted proteolysis. Interaction between the unfoldase, ClpX, and the substrate with an accessible degron initiates energy-dependent proteolysis. Four anti-{sigma} factors with a mutation in the degron provided a set of natural substrates to evaluate the influence of the degron on degradation strength in ClpX-substrate processivity. We note that a point mutation in the degron (XXX-Ala-Ala) leads to an order of magnitude difference in the dwell time of the substrate on ClpX. Differences in ClpX/anti-{sigma} interactions were correlated with change in unfoldase activity. GFP chimeras or polypeptides of identical length with the anti-{sigma} degron also demonstrate degron-dependent variation in ClpX activity. We show that degron-dependent ClpX activity leads to differences in anti-{sigma} factor degradation thereby regulating the release of free {sigma} from the {sigma}/anti-{sigma} complex. M. tuberculosis ClpX activity thus influences changes in gene expression by modulating the cellular abundance of ECF {sigma} factors.\n\nImportanceThe ability of Mycobacterium tuberculosis to quickly adapt to the changing environmental stimuli occurs by maintaining protein homeostasis. Extra-cytoplasmic function (ECF) {sigma} factors play a significant role in coordinating the transcription profile to changes in environmental conditions. Release of the {sigma} factor from the anti-{sigma} is governed by the ClpXP2P1 assembly. M. tuberculosis ECF anti-{sigma} factors have a ssrA-like degron for targeted degradation. A point mutation in the degron leads to differences in ClpX mediated proteolysis and affects the cellular abundance of ECF {sigma}-factors. ClpX activity thus synchronizes changes in gene expression with environmental stimuli affecting M. tuberculosis physiology.