Overexpression of the third H-NS paralogue H-NS2 compensates fitness loss in hns mutants of the enteroaggregative Escherichia coli strain 042

A Prieto,T Chakraborty,L Falgenhauer,M Bernabeu,A Juárez,M Hüttener

doi:10.1038/s41598-020-75204-4

Abstract

Members of the H-NS protein family play a role both in the chromosome architecture and in the regulation of gene expression in bacteria. The genomes of the enterobacteria encode an H-NS paralogue, the StpA protein. StpA displays specific regulatory properties and provides a molecular backup for H-NS. Some enterobacteria also encode third H-NS paralogues. This is the case of the enteroaggregative E. coli (EAEC) strain 042, which encodes the hns, stpA and hns2 genes. We provide in this paper novel information about the role of the H-NS2 protein in strain 042. A C > T transition in the hns2 promoter leading to increased H-NS2 expression is readily selected in hns mutants. Increased H-NS2 expression partially compensates for H-NS loss. H-NS2 levels are critical for the strain 042 fitness. Under some circumstances, high H-NS2 expression levels dictated by the mutant hns2 promoter can be deleterious. The selection of T > C revertants or of clones harboring insertional inactivations of the hns2 gene can then occur. Temperature also plays a relevant role in the H-NS2 regulatory activity. At 37 °C, H-NS2 targets a subset of the H-NS repressed genes contributing to their silencing. When temperature drops to 25 °C, the repressory ability of H-NS2 is significantly reduced. At low temperature, H-NS plays the main repressory role.

Highlights

Nucleoid-associated proteins of the H-NS family contribute to shaping the chromosome and regulate gene expression mainly in the enterobacteria and in other Gram-negative microorganisms
An experimental evolution approach designed to identify secondary mutations compensating for H-NS loss in Salmonella showed that, when subculturing Δhns mutants, some of the Δhns cell lineages that increased the growth rate accumulated mutations in the stpA gene, generating StpA variants with altered DNA binding and oligomerization properties, resembling those of H-NS29
We considered that a similar approach could be performed with strain 042 in order to gain insight into H-NS2 function

Summary

Introduction

Nucleoid-associated proteins of the H-NS family contribute to shaping the chromosome and regulate gene expression mainly in the enterobacteria and in other Gram-negative microorganisms. H-NS binds preferentially ATrich DNA stretches, and it has been proposed to operate as a xenogeneic silencer, avoiding unwanted expression of horizontally acquired g enes[7,8,9,10,11]. This protein modulates the expression of HGT DNA, and of core genes[12]. Some pathogenic E. coli strains can encode additional hns paralogues. The H-NS paralogues StpA, Sfh and Hfp have been proposed to modulate H-NS activity by forming heteromers with H-NS, which display DNA-binding properties different from those from homomeric H-NS25–27. We show a differential regulatory role of the H-NS2 protein at 25 °C and 37 °C

Methods

Results

Conclusion