Lsr2 of Mycobacterium Represents a Novel Class of H-NS-Like Proteins

Abstract

Lsr2 is a small, basic protein present in Mycobacterium and related actinomycetes. Our previous in vitro biochemical studies showed that Lsr2 is a DNA-bridging protein, a property shared by H-NS-like proteins in gram-negative bacteria. Here we present in vivo evidence based on genetic complementation experiments that Lsr2 is a functional analog of H-NS, the first such protein identified in gram-positive bacteria. We show that lsr2 can complement the phenotypes related to hns mutations in Escherichia coli, including ␤-glucoside utilization, mucoidy, motility, and hemolytic activity. We also show that Lsr2 binds specifically to H-NSregulated genes and the repression of hlyE by Lsr2 can be partially eliminated by overexpression of slyA, suggesting that the molecular mechanisms of Lsr2 repression and depression are similar to those of H-NS. The functional equivalence of these two proteins is further supported by the ability of hns to complement the lsr2 phenotype in Mycobacterium smegmatis. Taken together, our results demonstrate unequivocally that Lsr2 is an H-NS-like protein.

Lsr2 is a nucleoid-associated protein that targets AT-rich sequences and virulence genes inMycobacterium tuberculosis

“…We recently demonstrated by in vitro biochemical experiments that, like H-NS, Lsr2 of Mycobacterium tuberculosis (M. tb) binds DNA in a sequence-independent manner and is capable of bridging distant DNA segments (19). Moreover, we showed through in vivo complementation assays that Lsr2 is a functional analog of H-NSspecifically, that lsr2 fully complements independent phenotypes associated with hns mutations in E. coli (15). These results suggest that Lsr2 may play a role in M. tb that is equivalent to that of H-NS.…”

“…4B), representing a unique mechanism of DNA recognition. These residues, particularly Arg84 and Arg97-Gly98-Arg99, are highly conserved among Lsr2 homologs (15).…”

“…Lsr2 is a small, basic protein that is highly conserved in mycobacteria and related actinomycetes (15). Previous studies showed that Lsr2 is involved in several cellular processes including cellwall lipid biosynthesis (16,17) and antibiotic resistance (18).…”

“…The functional similarities between Lsr2 and H-NS could not be predicted from their primary sequence because Lsr2 exhibits <20% sequence identity with H-NS and has a different predicted secondary structure (15). H-NS has two functional domains: an N-terminal dimerization domain and a C-terminal DNA-binding domain connected by an unstructured linker (2).…”

“…Less is known about Lsr2, but it has been shown to form dimers in vivo (16). To map the functional domains of Lsr2, six chimeric protein constructs containing different combinations of the N-and C-terminal sequences of Lsr2 and H-NS were generated, and their ability to complement phenotypes associated with hns mutations in E. coli and lsr2 mutations in M. smegmatis was tested (15). Three constructs (C1, C4, and C6) could complement several phenotypes in hns mutants but were ineffective at complementing an lsr2 mutation in M. smegmatis (Table 1).…”