Development and Application of Two Inducible Expression Systems for Streptococcus suis

Abstract

ABSTRACT Streptococcus suis is an important zoonotic bacterial pathogen posing a threat to the pig industry as well as public health, for which the mechanisms of growth and cell division remain largely unknown. Developing convenient genetic tools that can achieve strictly controlled gene expression is of great value for investigating these fundamental physiological processes of S. suis. In this study, we first identified three strong constitutive promoters, Pg, Pt, and Pe, in S. suis. Promoter Pg was used to drive the expression of repressor genes tetR and lacI, and the operator sequences were added within promoters Pt and Pe. By optimizing the insertion sites of the operator sequence, we successfully constructed an anhydrotetracycline (ATc)-inducible expression system and an isopropyl-β-d-thiogalactopyranoside (IPTG)-inducible expression system in S. suis. We showed that these two systems provided inducer-concentration- and induction-time-dependent expression of the reporter gene. By using these tools, we investigated the subcellular localization of a key cell division protein, FtsZ, which showed that it could be correctly localized to the midcell region. In addition, we constructed a conditional knockout strain for the glmS gene, which is an essential gene, and showed that our ATc-inducible promoter could provide strictly controlled expression of glmS in trans, suggesting that our inducible expression systems can be used for deletion of essential genes in S. suis. Therefore, for the first time we developed two inducible expression systems in S. suis and showed their applications in the study of an important cell division protein and an essential gene. These genetic tools will further facilitate the functional study of other important genes of S. suis. IMPORTANCE Streptococcus suis is an important zoonotic bacterial pathogen. Studying the mechanisms of cell growth and division is important for the identification of novel antimicrobial drug targets. Inducible expression systems can provide strictly controlled expression of the protein of interest and are useful tools to study the functions of physiologically important proteins. However, there is a lack of convenient genetic tools that can achieve inducible protein expression in S. suis. In this study, we developed two (ATc-inducible and IPTG-inducible) inducible expression systems and showed their applications in a subcellular localization study of a cell division protein and the construction of conditional knockout of essential genes in S. suis. These systems will be useful for functional studies of important proteins of S. suis.

Keywords

• Streptococcus suis
• genetic tool
• inducible expression system
• subcellular localization
• conditional gene knockout