Multiple Mechanisms Are Involved in Repression of Filamentous Phage SW1 Transcription by the DNA-Binding Protein FpsR

Huahua Jian; Guanpeng Xu; Shunzhang Liu; Yali Hao; Canxing Meng; Jianrong Xu; Yue Zhang; Xipeng Liu; Xiang Xiao

doi:10.1016/j.jmb.2019.01.040

Multiple Mechanisms Are Involved in Repression of Filamentous Phage SW1 Transcription by the DNA-Binding Protein FpsR

J Mol Biol. 2019 Mar 15;431(6):1113-1126. doi: 10.1016/j.jmb.2019.01.040. Epub 2019 Feb 7.

Authors

Huahua Jian¹, Guanpeng Xu², Shunzhang Liu², Yali Hao², Canxing Meng², Jianrong Xu³, Yue Zhang², Xipeng Liu², Xiang Xiao⁴

Affiliations

¹ State Key Laboratory of Microbial Metabolism, School of Life Sciences and Biotechnology, Shanghai Jiao Tong University, Shanghai 200240, PR China; State Key Laboratory of Ocean Engineering, School of Naval Architecture, Ocean and Civil Engineering, Shanghai Jiao Tong University, Shanghai 200240, PR China; Laboratory for Marine Biology and Biotechnology, Qingdao National Laboratory for Marine Science and Technology, Qingdao 266237, PR China.
² State Key Laboratory of Microbial Metabolism, School of Life Sciences and Biotechnology, Shanghai Jiao Tong University, Shanghai 200240, PR China.
³ Department of Pharmacology and Chemical Biology, Institute of Medical Sciences, School of Medicine, Shanghai Jiao Tong University, Shanghai 200025, PR China.
⁴ State Key Laboratory of Microbial Metabolism, School of Life Sciences and Biotechnology, Shanghai Jiao Tong University, Shanghai 200240, PR China; State Key Laboratory of Ocean Engineering, School of Naval Architecture, Ocean and Civil Engineering, Shanghai Jiao Tong University, Shanghai 200240, PR China; Laboratory for Marine Biology and Biotechnology, Qingdao National Laboratory for Marine Science and Technology, Qingdao 266237, PR China. Electronic address: zjxiao2018@sjtu.edu.cn.

PMID: 30738893
DOI: 10.1016/j.jmb.2019.01.040

Abstract

SW1 is the first filamentous phage isolated from a deep-sea environment. Nevertheless, the mechanism by which the SW1 genetic switch is controlled is largely unknown. In this study, the function of the phage-encoded FpsR protein was characterized by molecular biological and biochemical analyses. The deletion of fpsR increased the copy number of SW1 ssDNA and mRNA, indicating that FpsR functions as a repressor. In addition, transcription from the fpsR promoter was shown to be increased in an fpsR deletion mutant, suggesting self-repression by FpsR. Purified FpsR bound to four adjacent operator sites (O1-O4) embedded within the fpsA promoter and the fpsA-fpsR intergenic region. A surface plasmon resonance experiment showed that FpsR can bind to the O1-O4 operators separately and with different binding affinity, and the dissociation constants of FpsR with O2 and O3 were found to be lower at 4 °C than at 20 °C. A gel permeation chromatography assay revealed that FpsR oligomerized to form tetramers. Point mutation analysis indicated that the C-terminal domain influenced the binding affinity and regulatory function of FpsR. Collectively, these data support a model in which FpsR actively regulates phage production by interacting with the corresponding operators, thus playing a crucial role in the SW1 genetic switch.

Keywords: FpsR; deep sea; filamentous phage; repressor; transcriptional repression.

Publication types

Research Support, Non-U.S. Gov't

MeSH terms

DNA Replication
DNA-Binding Proteins / genetics
DNA-Binding Proteins / metabolism*
Gene Expression Regulation, Viral*
Inovirus / genetics*
Inovirus / metabolism
RNA, Viral / genetics*
Transcriptional Activation
Viral Proteins / genetics
Viral Proteins / metabolism*

Substances

DNA-Binding Proteins
RNA, Viral
Viral Proteins