Control of Mycobacterium tuberculosis protein secretion by ESX-4 and the outer membrane EsxUT-EsxEF complex

Mycobacterium tuberculosis uses five ESX systems to secrete effector proteins with distinct functions essential for the pathogen's growth and virulence. The current paradigm assumes that each ESX system secretes a distinct set of effector proteins. Here, we show that the EsxU-EsxT proteins, associated with the ESX-4 system, and the EsxE-EsxF proteins, encoded in the toxin operon cpnT, form a supercomplex that is required not only for toxin secretion, but also for outer membrane localization and for secretion of all Esx proteins into the cytosol of infected macrophages. Secretion of non-Esx proteins such as EspB, proteins involved in iron uptake, and PPE62, which are dependent on ESX-1, ESX-3 and ESX-5, respectively, also requires the EsxUT-EsxEF supercomplex. The mutual functional dependency of EsxEF and EsxUT on each other for outer-membrane localization synchronizes effector protein secretion, enabling M. tuberculosis to block phagosomal maturation and to permeabilize the phagosomal membrane when the pathogen is capable of killing host cells by toxin secretion. The requirement of the ESX-4 system for general secretion of ESX-dependent proteins in vivo is a critical vulnerability that could be targeted by drugs and/or in vaccines to simultaneously block many virulence factors of M. tuberculosis.