Host DHFRL1 interacts with MtbEis and increases the intracellular survival of Mycobacterium tuberculosis by inhibiting autophagy.

The incredible ability of Mycobacterium tuberculosis to orchestrate the host cell machinery is achieved by its secretory proteins. M. tuberculosisEnhanced Intracellular Survival (MtbEis) protein is one such virulence-associated secretory acetyltransferase that acetylates aminoglycoside antibiotics, arylalkylamines, mycobacterial proteins and host proteins. To reveal the unknown functions of MtbEis, we used Yeast Two-Hybrid assay to explore its interactions with host proteins. We discovered a novel interaction of MtbEis with mouse Dihydrofolate Reductase (mDHFR) protein and demonstrated that MtbEis specifically acetylates mDHFR at K158 and K177 residues. Additionally, we confirmed the interaction of MtbEis with endogenous human DHFR. MtbEis localizes to mitochondria in addition to the cytoplasm and interacts with the mitochondrial isoform of human DHFR, known as DHFR like 1 (DHFRL1). We observed a distinct pattern of mRNA expression for hDHFR and DHFRL1 in M. tuberculosis-infected THP1 macrophages. Using siRNA, inhibition of DHFRL1 expression but not cytoplasmic DHFR (hDHFR) in THP1 macrophages significantly reduced the intracellular survival of M. tuberculosis. Notably, DHFRL1 knockdown led to enhanced autophagy activation and increased lysosomal delivery of M. tuberculosis in host macrophages. In summary, our study identifies a novel post-translational modification of mouse DHFR and unravels a previously unrecognized role of human DHFRL1 protein in supporting the intracellular survival of M. tuberculosis. Also, these findings highlight distinct functional contributions of cytosolic and mitochondrial isoforms of human DHFR in pathogen biology.