Lysosome repositioning as an autophagy escape mechanism by Mycobacterium tuberculosis Beijing strain

Abstract

Induction of host cell autophagy by starvation was shown to enhance lysosomal delivery to mycobacterial phagosomes, resulting in the restriction of Mycobacterium tuberculosis reference strain H37Rv. Our previous study showed that strains belonging to M. tuberculosis Beijing genotype resisted starvation-induced autophagic elimination but the factors involved remained unclear. Here, we conducted RNA-Seq of macrophages infected with the autophagy-resistant Beijing strain (BJN) compared to macrophages infected with H37Rv upon autophagy induction by starvation. Results identified several genes uniquely upregulated in BJN-infected macrophages but not in H37Rv-infected cells, including those encoding Kxd1 and Plekhm2, which function in lysosome positioning towards the cell periphery. Unlike H37Rv, BJN suppressed enhanced lysosome positioning towards the perinuclear region and lysosomal delivery to its phagosome upon autophagy induction by starvation, while depletion of Kxd1 and Plekhm2 reverted such effects, resulting in restriction of BJN intracellular survival upon autophagy induction by starvation. Taken together, these data indicated that Kxd1 and Plekhm2 are important for the BJN strain to suppress lysosome positioning towards the perinuclear region and lysosomal delivery into its phagosome during autophagy induction by starvation to evade starvation-induced autophagic restriction.

The autophagy-resistant Mycobacterium tuberculosis Beijing strain upregulates KatG to evade starvation-induced autophagic restriction

“… 2015 ; Laopanupong et al . 2021 ), enhanced colocalization of H37Rv with Cathepsin D was observed upon autophagy induction by the starvation of Raw264.7 macrophages, while such effect was not seen with the BJN (Fig. 2A and B ).…”

“… 2015 ; Laopanupong et al . 2021 ), starvation-induced autophagy of RAW264.7 macrophages resulted in the restriction of M. tuberculosis reference strain H37Rv, while the BJN resisted such elimination (Fig. 1B ).…”

“…In addition, by using RNA-Seq analysis of H37Rv- and BJN-infected macrophages, we recently identified host factors, Kxd1 and Plekhm2 which function in lysosome positioning toward the cell periphery, to contribute to autophagy resistance by the BJN (Laopanupong et al . 2021 ). By upregulating the aforementioned host proteins in the infected macrophages, the BJN could promote the lysosome distribution toward the cell periphery and away from its phagosome, thereby sparing itself from autophagy-mediated lysosomal delivery and restriction (Laopanupong et al .…”

“… 2015 ; Laopanupong et al . 2021 ) were propagated in Middlebrook 7H9 broth or on 7H10 agar supplemented with 10% oleic acid–albumin–dextrose–catalase (OADC; BD), 0.05% Tween 80, and 0.2% glycerol at 37°C. Before the experiments, logarithmic phase cultures were collected, washed twice with PBS, and resuspended in the full medium.…”

“…We also recently showed that, through RNA-Seq analysis of the infected host cells, macrophages infected with the BJN upregulated gene function in lysosome positioning toward cell periphery to facilitate the escape of the BJN from starvation-induced autophagic restriction (Laopanupong et al . 2021 ). However, the mycobacterial factors involved in the autophagy resistance by the BJN remain unknown.…”