WhiB6 is required for the secretion-dependent regulation of ESX-1 substrates in pathogenic mycobacteria

Abstract

23The mycobacterial type VII secretion system ESX-1 is responsible for secretion of a number 24 of proteins that play an important role during infection of the host. Regulation of expression 25 of secreted proteins is often essential to establish a successful infection. Using transcriptome 26 sequencing, we found that abrogation of ESX-1 function in Mycobacterium marinum leads to 27 a pronounced increase in gene expression levels of the espA operon during infection of 28 macrophages, suggesting an important role in ESX-1-mediated virulence during the early 29 phase of infection. In addition, we found that disruption of ESX-1-mediated protein secretion 30 leads to a specific down-regulation of the substrates, but not the structural components of this 31 system, in both M. marinum and M. tuberculosis during growth in culture medium. We 32 established that down-regulation of ESX-1 substrates is the result of a regulatory process, 33 which is influenced by the putative transcriptional regulator whib6, located adjacent to the 34 esx-1 locus. In addition, overexpression of the ESX-1 associated PE35/PPE68 protein pair 35 resulted in significantly increased secretion of the ESX-1 substrate EsxA, demonstrating a 36 functional link between these proteins. Together, these data show that ESX-1 substrates are 37 regulated independently from the structural components, both during infection and as a result 38 of active secretion. 39 40 3