Influence of SmpB and ClpX interactions and interactomes on the transcription profile of Mycobacterium tuberculosis

SmpB is a key regulator of trans-translation in bacteria. The interactome of Mycobacterium tuberculosis (Mtb) ClpX revealed SmpB alongside multiple transcriptional regulators and sigma factors that can alter the transcriptional profile. The finding that ClpX could also be identified in the Mtb SmpB interactome suggested both physical and functional linkages between trans-translation and targeted protein degradation. The trans-translation mechanism facilitates the release of stalled ribosomes and the attachment of ssrA degron sequences to aberrant proteins. We show that both SmpB and the previously described Mtb ClpX adaptor, SSB, bind to the N-terminal domain of ClpX. However, unlike the adaptor SSB, SmpB binding does not enhance ClpX ATPase activity. Interaction studies using a model substrate with an exposed ssrA degron revealed that SmpB enhances ClpX-substrate interactions. ClpX unfoldase activity, on the other hand, enables targeted degradation of proteins containing the ssrA degron. ClpX also facilitates the release of specific s factors from inactive sigma/anti-sigma complexes by degrading anti-sigma factors containing the ssrA degron thus indirectly influencing the transcription profile. Together, these observations suggest that SmpB and ClpX interactions and interacting protein networks exert both direct as well as indirect effects on the Mtb transcriptional profile.