Oxygen sensing and transcriptional regulation under hypoxia exposure in the mollusk Crassostrea gigas.

Abstract

Hypoxia caused by global climate change and anthropogenic pollution has exposed marine species to increasing stress. Oxygen sensing mediated by prolyl hydroxylase (PHD) is regarded as the first line of defense under hypoxia exposure; however, the function of PHD in marine molluscan species remains unclear. In this study, we identified two PHD2 gene in the oyster Crassostrea gigas using phylogenetic tree analysis with 36 species, namely, CgPHD2A/B. Under hypoxia, the mRNA and protein expression of CgPHD2A displayed a time-dependent pattern, revealing a critical role in the response to hypoxia-induced stress. Observation of interactions between CgPHD2 and CgHIF-1α proteins under normoxia using co-immunoprecipitation and GST-pull down experiments showed that the β2β3 loop in CgPHD2A hydroxylates CgHIF-1α to promote its ubiquitination with CgVHL. With the protein recombination and site-directed mutagenesis, the hydroxylation domain and two target proline loci (P404A and 504A) in CgPHDs and CgHIF-1α were identified respectively. Moreover, the electrophoretic mobility-shift assay (EMSA) and luciferase double reporter gene assay revelaed that CgHIF-1α could regulate CgPHD2A expression through binding with the hypoxia-responsive element in the promoter region (320 bp upstream), forming a feedback loop. However, protein structure analysis indicated that six extra amino acids formed an α-helix in the β2β3 loop of CgPHD2B, inhibiting its activity. Overall, this study revealed that two CgPHD2 proteins have evolved, which encode enzymes with different activities in oyster, potentially representing a specific hypoxia-sensing mechanism in mollusks. Illustrating the functional diversity of CgPHDs could help to assess the physiological status of oyster and guide their aquaculture.

• Authors: Jie Meng, Ting Wang, Busu Li, Li Li, Guofan Zhang
• Journal: The Science of the total environment (Sci Total Environ) Vol. 853 Pg. 158557 (Dec 20 2022) ISSN: 1879-1026 [Electronic] Netherlands
• PMID: 36084780 (Publication Type: Journal Article)
• Copyright: Copyright © 2022. Published by Elsevier B.V.

Chemical References

• Oxygen
• Procollagen-Proline Dioxygenase
• RNA, Messenger
• Proline
• Amino Acids

Topics

• Animals
• Crassostrea (genetics, metabolism)
• Oxygen (metabolism)
• Phylogeny
• Hypoxia
• Procollagen-Proline Dioxygenase (genetics, metabolism)
• RNA, Messenger (genetics)
• Proline (genetics)
• Amino Acids