Cloning and characterization of Rap GTPase from the Chinese white shrimp Fenneropenaeus chinensis

Abstract

Ras-related protein Rap GTPase has been implicated in cell adhesion, cell proliferation, and cell junction formation. The first shrimp Rap cDNA ( FcRap) was recently identified from the Chinese white shrimp Fenneropenaeus chinensis. The full length of FcRap is 1013 bp, with a 561 bp open reading frame that encodes a 186 amino acid protein. FcRap has a calculated molecular mass of 20.90 kDa and p I of 6.37. Phylogenetic analysis shows that FcRap and other Rap proteins are clustered into one group. Results from the quantitative real-time polymerase chain reaction show that FcRap could be detected mainly in the hemocytes, hepatopancreas, stomach, and gills, whereas a relatively lower expression level could be detected in the heart and intestines. FcRap in the hemocytes was upregulated 2 h post Vibrio challenge, and it was upregulated 2 h post white spot syndrome virus (WSSV) challenge, and peaked at 6 h before it declined at 12 h. No variation in the FcRap transcript was observed in the gills under the Vibrio challenge, but it was initially downregulated 2 h post WSSV challenge, and then it was upregulated and peaked at 6 h before it was eventually went down at 12 h. The rFcRap protein was successfully expressed in Escherichia coli BL21DE3. The pull-down analysis showed that rFcRap protein could interact with VP28, an envelope protein of WSSV. The probable roles of Rap GTPase in shrimp innate immunity are presented for the first time.