The SapM phosphatase arrests phagosome maturation in an ESX-1 independent manner inMycobacterium tuberculosisand BCG

AbstractMycobacterium tuberculosis(Mtb) is an intracellular pathogen that survives and grows in macrophages. A mechanism used byMtbto achieve intracellular survival is to secrete effector molecules that arrest the normal process of phagosome maturation. Through phagosome maturation arrest (PMA),Mtbremains in an early phagosome and avoids delivery to degradative phagolysosomes. One PMA effector ofMtbis the secreted SapM phosphatase. Because the host target of SapM, phosphatidylinositol-3-phosphate (PI3P), is located on the cytosolic face of the phagosome, SapM needs to be both released by the mycobacteria and escape the phagosome to carry out its function. To date, the only mechanism known forMtbmolecules to escape the phagosome is phagosome permeabilization by the ESX-1 secretion system. To understand this step of SapM function in PMA, we generated identical in-framesapMmutants in both the attenuatedMycobacterium bovisbacille Calmette-Guérin (BCG) vaccine strain, which lacks the ESX-1 system, andMtb. Characterization of these mutants demonstrated that SapM is required for PMA in both BCG andMtb. Further, by establishing a role for SapM in PMA in BCG, and subsequently in aMtbmutant lacking the ESX-1 system, we demonstrated that the role of SapM is ESX-1-independent. We further determined that ESX-2 or ESX-4 are also not required for SapM to function in PMA. These results indicate that SapM is a secreted effector of PMA in both BCG andMtband that it functions independent of the known mechanism forMtbmolecules to escape the phagosome.