Cloning and Expression Characteristics Analysis of the Aquaporin Gene EHP00_492 from Enterocytozoon hepatopenaei

Chenxi LI; Huixin WU; Yujiao WU; Jie CHEN; Xianzhi MENG; Guoqing PAN; Mengxian LONG

doi:10.16768/j.issn.1004-874X.2023.08.016

Guangdong nongye kexue (Aug 2023)

Cloning and Expression Characteristics Analysis of the Aquaporin Gene EHP00_492 from Enterocytozoon hepatopenaei

Chenxi LI,
Huixin WU,
Yujiao WU,
Jie CHEN,
Xianzhi MENG,
Guoqing PAN,
Mengxian LONG

Affiliations

Chenxi LI: State Key Laboratory of Resource Insects/Chongqing Key Laboratory of Microsporidia Infection and Control, Southwest University, Chongqing 400715, China
Huixin WU: State Key Laboratory of Resource Insects/Chongqing Key Laboratory of Microsporidia Infection and Control, Southwest University, Chongqing 400715, China
Yujiao WU: State Key Laboratory of Resource Insects/Chongqing Key Laboratory of Microsporidia Infection and Control, Southwest University, Chongqing 400715, China
Jie CHEN: State Key Laboratory of Resource Insects/Chongqing Key Laboratory of Microsporidia Infection and Control, Southwest University, Chongqing 400715, China
Xianzhi MENG: State Key Laboratory of Resource Insects/Chongqing Key Laboratory of Microsporidia Infection and Control, Southwest University, Chongqing 400715, China
Guoqing PAN: State Key Laboratory of Resource Insects/Chongqing Key Laboratory of Microsporidia Infection and Control, Southwest University, Chongqing 400715, China
Mengxian LONG: State Key Laboratory of Resource Insects/Chongqing Key Laboratory of Microsporidia Infection and Control, Southwest University, Chongqing 400715, China

DOI: https://doi.org/10.16768/j.issn.1004-874X.2023.08.016
Journal volume & issue: Vol. 50, no. 8
pp. 154 – 162

Abstract

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【Objective】The aquaporin gene EHP00_492 from Enterocytozoon hepatopenaei (EHP) was cloned and its expression characteristics in EHP mature spores was analyzed, which provided a theoretical basis for studying the function of EHP aquaporin.【Method】The complete EHP00_492 gene was cloned from Ent. hepatopenaei. The sequence characteristics of EHP00_492 was analyzed by bioinformatics methods. Recombinant expression vector pCold-TF-EHP00_492 was constructed and transformed into Escherichia coli to induced fusion protein expression for preparing the rabbit polyclonal antibody of EHP00_492. The expression and subcellular localization characteristics of EHP00_492 in EHP mature spores were analyzed by Western blot and indirect immunofluorescence assay.【Result】EHP00_492 gene is 729 bp in length, encoded 242 amino acids. EHP00_492 is rich in leucine, with a predicted molecular weight of 25 kD. It has no signal peptide, but involving six transmembrane domains, the MIP conserved domain and two aquaporin conserved NPA/G motif. In addition, EHP00_492 has a high homology with other microsporidian aquaporins, and the phylogenetic tree analysis result shows that EHP00_492 is closely related with EBI_27080 protein from Ent. bieneusi. The three-dimensional structure of EHP00_492 is highly similarity with the identified aquaporin NbAQP from Nosema bombycis and EcAQP from Enc. cuniculi by AlphaFold analysis, which is speculated that EHP00_492 is a potential aquaporin. EHP00_492 is a 21 kD-protein in the mature spores of EHP by Western blot analysis. And the subcellular localization analysis showed that EHP00_492 protein was localized at the spore wall of EHP mature spores.【Conclusion】In this study, the sequence and structure characteristics of EHP00_492 protein, as well as its phylogenetic relationship with other microsporidian aquaporins, and the expression and subcellular localization characteristics of EHP00_492 protein in EHP mature spores were preliminary clarified, which provided basis for further study on the function of EHP aquaporins.

Published in Guangdong nongye kexue

ISSN: 1004-874X (Print)
Publisher: Guangdong Academy of Agricultural Sciences
Country of publisher: China
LCC subjects: Agriculture
Website: http://gdnykx.cnjournals.org/gdnykx/ch/index.aspx

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