Structural analysis of the HicAB toxin-antitoxin system encoded by a prophage in coral-associated Halomonas meridiana

  • CHEN Ran
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  • 1 College of Life Science and Technology, Jinan University, Guangzhou, Guangdong, 510632, China

    2 Key Laboratory of Tropical Marine Bio-resources and Ecology, South China Sea Institute of Oceanology, Chinese Academy of Sciences, Guangzhou 511458, China

Received date: 2025-02-17

  Revised date: 2025-03-14

  Accepted date: 2025-03-18

  Online published: 2025-03-18

Supported by

National Science Foundation of China(42188102); Special Fund of the South China Sea Institute of Oceanology, Chinese Academy of Sciences(SCSIO2023QY03); the Ocean Negative Carbon Emissions (ONCE) Program; Open project of State Key Laboratory of South China Sea Marine Resources Utilization, Hainan University(MRUKF2023001)

Abstract

The interaction between phages and bacterial hosts significantly impacts coral health and reef stability, representing a key focus in virology. To combat phage infection, bacteria have evolved diverse innate and adaptive immune systems, including toxin-antitoxin (TA) systems, a crucial defense mechanism. In this study, bioinformatics analysis predicted that the CTT34_05265 and CTT34_05260 genes within the prophage Phm2 of the coral-associated bacterium Halomonas meridiana SCSIO 43005 (Hm43005) encode HmHicA and HmHicB proteins, respectively. Their TA system functionality was confirmed through E. coli growth assays. Pull-down and bacterial two-hybrid experiments validated the interaction between HmHicA and HmHicB. Electrophoretic mobility shift assays (EMSA) and DNase I footprinting revealed that HmHicB binds specifically to palindromic sequences upstream of the -35 and -10 regions of the HmhicA promoter, mediating transcriptional autoregulation. Additionally, HmHicB alone exhibited mild toxicity, suggesting potential alternative regulatory targets for the antitoxin. Structural analysis indicated that HmHicA functions as a ribosome-independent RNase, while HmHicB contains an N-terminal toxin-binding domain and a C-terminal DNA-binding domain. HmHicB forms a homodimer via its C-terminus and assembles with HmHicA in a 2:2 stoichiometric complex. The molecular mechanism of antitoxin-mediated toxin inhibition likely involves electrostatic interactions between the positively charged active pocket of HmHicA and the negatively charged toxin-binding domain of HmHicB, as well as the burial of the HmHicA His24 active site. These findings provide a foundation for further exploration of the properties and physiological roles of this TA system.

Cite this article

CHEN Ran . Structural analysis of the HicAB toxin-antitoxin system encoded by a prophage in coral-associated Halomonas meridiana[J]. Journal of Tropical Oceanography, 0 : 1 . DOI: 10.11978/2025022

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