Gene Regulatory and Physiological Functions of Histone-Like Protein from Streptococci

Objectives: Streptococcal histone-like DNA binding protein is a homodimeric protein and conserved with Escherichia coli HU, a well-documented nucleoid-associated protein. In E. coli, HU plays important roles as both structural and regulatory factors, but it is not essential for E. coli viability. HLP from Streptococcus intermedius (Si-HLP) has been found to bind host cells and induce pro-inflammatory cytokine production, but its physiological role remains poorly defined. The aims of this study were to investigate the essentiality of Si-HLP for bacterial viability and normal growth and to examine the structural and gene regulatory functions of Si-HLP.

Methods: Using gene insertion knockout and tetracycline-regulated antisense RNA expression techniques (inducible antisense RNA expression system), we determined whether Si-HLP is essential for bacterial viability and normal growth in S. intermedius. In wild-type (WT) and Si-HLP-down-regulated strain, we also investigated the morphological change and cell surface characteristic by electron microscopic observation and cell surface hydrophobicity, respectively. Moreover, nucleoid compactness and gene regulatory functions of Si-HLP were examined by DNA staining, real-time PCR and RT-PCR in WT and Si-HLP-down-regulated strain.

Results: Tetracycline-induced Si-hlp antisense RNA expression specifically inhibited endogenous Si-HLP protein expression and the inhibitory growth of the Si-HLP-down-regulated mutant by this inducible antisense mRNA technique was observed. The Si-HLP-down-regulated strain showed alterations in its cell morphology such as cell size, chain formation and cell divisions, as well as cell surface hydrophobicity. Down-regulation of Si-HLP led to an expanded nucleoid to fill the intracellular space. Transcription levels of several genes, including virulence-associated factors, were found to be activated or repressed in the antisense Si-hlp RNA-expressing strain.

Conclusion: These data demonstrated that Streptococcal HLP is essential for cell viability and normal growth in Streptococci and suggest that Streptococcal HLP serves as an essential nucleoid-associated protein governing the nucleoid architecture and controlling the gene transcription profile in Streptococci.