Lsr2, a nucleoid-associated protein influencing mycobacterial cell cycle

Abstract

Nucleoid-associated proteins (NAPs) are responsible for maintaining highly organized and yet dynamic chromosome structure in bacteria. The genus Mycobacterium possesses a unique set of NAPs, including Lsr2, which is a DNA-bridging protein. Importantly, Lsr2 is essential for the M. tuberculosis during infection exhibiting pleiotropic activities including regulation of gene expression (mainly as a repressor). Here, we report that deletion of lsr2 gene profoundly impacts the cell morphology of M. smegmatis, which is a model organism for studying the cell biology of M. tuberculosis and other mycobacterial pathogens. Cells lacking Lsr2 are shorter, wider, and more rigid than the wild-type cells. Using time-lapse fluorescent microscopy, we showed that fluorescently tagged Lsr2 forms large and dynamic nucleoprotein complexes, and that the N-terminal oligomerization domain of Lsr2 is indispensable for the formation of nucleoprotein complexes in vivo. Moreover, lsr2 deletion exerts a significant effect on the replication time and replisome dynamics. Thus, we propose that the Lsr2 nucleoprotein complexes may contribute to maintaining the proper organization of the newly synthesized DNA and therefore influencing mycobacterial cell cycle.

Lsr2 and Its Novel Paralogue Mediate the Adjustment of Mycobacterium smegmatis to Unfavorable Environmental Conditions

“…This is in line with the subcellular localization of Lsr2, which is not distributed throughout the nucleoid but rather forms one or two nucleoprotein complexes (Fig. 5C) (23). We speculate that changes in the LOS level, which were presumably mirrored by differences in the cell envelope properties of Dlsr2 cells, may have directly altered the penetration of antibiotics (45).…”

“…3B and 5C), while Lsr2 forms a highly dynamic nucleoprotein complex (or two complexes), as we recently demonstrated (Fig. 5C) (23). Thus, the subcellular localizations of MSMEG_1060 and Lsr2 reflect their DNA binding preferences.…”

“…Thus, the localization of MSMEG_1060-mTurquoise2 substantially differed from that of Lsr2, which was visible as one or two discrete and bright major foci (see Fig. 5C) (23).…”

“…Notably, the majority of the Lsr2 binding sites identified by our ChIP-seq analysis (532 sites; 91%) are not directly (or at all) associated with transcriptional regulation. These sites may play an architectural role in the formation of massive nucleoprotein Lsr2-DNA complexes that presumably contribute to maintaining the proper organization of the newly synthesized DNA (23). However, we cannot exclude the possibility that Lsr2-induced changes in DNA topology and/or DNA accessibility may indirectly affect the expression levels of some genes, particularly under unfavorable conditions (e.g., oxygen depletion [see below]).…”

“…DNA manipulations, bacterial strains, and culture conditions. Bacterial strain construction and culture were performed as described previously (23). A detailed description of the bacterial culture conditions is provided in the Text S1 in the supplemental material.…”