Fig 1. Mutants affected in the ESX-1 accessory proteins EspG1, EspH and EccA1 differently affect the ESX-1 secretome.
A. Genetic organization of espG1-espH-eccA1 in the esx-1 locus. Genes are color-coded according to the localization of their proteins—see key. B and C. Secretion analysis of EsxA and EspE substrates reveals that single deletion of espG1, espH and eccA1 affects secretion at different levels. Immunoblot analysis using protein preparations of wild-type M. marinum and the indicated mutants. In B we analyzed cell pellets not treated with detergent Genapol X-080 and culture supernatant fractions. In C we analyzed cell pellets treated with Genapol X-080 and the concomitant supernatant fractions. D and E. Complementation of the mutant strains fully restores ESX-1 secretion. In D the secretion of EsxA was analyzed and in E the secretion of EspE. In both experiments, GroEL2 was used as loading control and PE_PGRS as cell-surface control fraction. Equivalent OD units were loaded; 0.2 OD for pellet or Genapol pellet and 0.5 OD for supernatant or Genapol supernatant fractions.