Role of hsp20 in the Production of Spores and Insecticidal Crystal Proteins in Bacillus thuringiensis.

Junyan Xie,Shuqing Yang,Xiaoli Zhao,Pengji Zhou,Liqiu Xia,Zixian Yi,Meifang Quan,Jiaoyang Lu,Tong Zhang,Jinli Peng,Shuiming Li,Xuezhi Ding

doi:10.3389/fmicb.2019.02059

Abstract

The small heat shock protein plays an important role in response to stresses. We wanted to investigate how Hsp20 affects sporulation and production of insecticidal crystal proteins (ICPs) in Bacillus thuringiensis (Bt) at the stationary growth phase when cells are starved. The hsp20 gene was knocked out in Bt4.0718 (wide type), which is a B. thuringiensis strain screened in our laboratory, using endonuclease I-SceI mediated unmarked gene replacement method. Deletion of Hsp20 resulted in a decrease in both sporulation and ICPs production. Bt4-Δhsp20 cells and its ICP did not have a significant difference in shape and size but entered the decline phase 2 h earlier than the Bt4.0718. In order to find the mechanism that underlies these phenotypes, we completed a proteomic study of differentially expressed proteins (DEPs). In Bt4-Δhsp20 cells, 11 DEPs were upregulated and 184 DEPs downregulated. These affected DEPs are involved in multiple metabolic pathways: (1) six DEPs (two upregulated and four downregulated) are directly related to the sporulation and ICPs synthesis; (2) supply of amino acids including amino acid synthesis and protein recycling; (3) the energy supplementation (the tricarboxylic acid cycle and glycolysis); (4) purine metabolism and mRNA stability. These results suggest that hsp20 may be critical in maintaining the homeostasis of B. thuringiensis during the production of spores and ICPs, and could provide new sight into the sporulation and ICPs formation in B. thuringiensis.

Highlights

Heat shock proteins (HSPs) are a family of proteins that are produced in response to stressful conditions, providing a chaperone function by stabilizing proteins to ensure correct folding, or by helping to refold proteins that are damaged by cell stress (Maio, 1999)
The expression of pET28a-hsp20 was induced in BL21(DE3) to prepare the polyclonal antibody, and whole proteins of the three strains were extracted for Western blot (WB) analysis verification (30S ribosome protein served as the internal reference)
The hsp20 would influence the production of sporulation and insecticidal crystal proteins (ICPs), which are the hallmarks of Bacillus thuringiensis (Bt), and the general metabolism

Summary

Introduction

Heat shock proteins (HSPs) are a family of proteins that are produced in response to stressful conditions, providing a chaperone function by stabilizing proteins to ensure correct folding, or by helping to refold proteins that are damaged by cell stress (Maio, 1999). The principal HSPs include five conserved classes, which are HSP33, HSP60, HSP70, HSP90, and HSP100, and small HSPs (sHSPs) (Schlesinger, 1990).

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