Cloning and characterization of Bax1 and Bax2 genes of Ctenopharyngodon idellus and evaluation of transcript expression in response to grass carp reovirus infection

Abstract

Multidomain proapoptotic Bcl-2-associated X (Bax) protein is an essential effector responsible for mitochondrial outer membrane permeabilization, resulting in cell death via apoptosis. In this study, two Bax genes of grass carp (Ctenopharyngodon idellus), designated as CiBax1 and CiBax2, were isolated and analyzed. The obtained CiBax1 cDNA is 2058 bp long, with a 579 bp open reading frame (ORF) coding a protein of 192 amino acid residues. The full-length cDNA of CiBax2 is 1161 bp, with a 618 bp ORF coding 205 amino acids. Both CiBax1 and CiBax2 are typical members of Bcl-2 family containing conserved Bcl and C-terminal domains, and they share conserved synteny with zebrafish Bax genes despite the grass carp Bax mapping to different linkage groups. Phylogenetic analysis showed that CiBax1 was clustered with Bax from most teleost fish, and CiBax2 was close to Bax2 from teleost fish but far separated from that of Salmo salar. Quantitative real-time PCR analysis revealed broad expression of CiBax1 and CiBax2 in tissues from healthy grass carp, but the relative expression level differed. The mRNA expression of CiBax1 and CiBax2 was both upregulated significantly and peaked in all examined tissues at days 5 or 6 post-infection with grass carp reovirus. Subcellular localization indicated that CiBax1 protein was localized in both nucleus and cytosol, while CiBax2 protein only in cytosol. Moreover, CiBax2, but not CiBax1 was colocalized with mitochondrion under normal condition. Taken together, the findings would be helpful for further understanding of the function of Bax in teleost fish.

Bid-deficient fish delay grass carp reovirus (GCRV) replication and attenuate GCRV-triggered apoptosis

“…Therefore, it is not surprising that the mRNA expression level of Bid was up-regulated at the late stage of infection. Similar expression pattern was also observed in other pro-apoptotic genes in grass carp [19,32]. Moreover, the dynamic changes of GCRV copy number in infected grass carp was detected in our previously study [33].…”

“…In addition, the expression profiles of CiCaspase-9 and CiCaspase-3 in liver and intestine after GCRV infection were also examined. As shown in Figure 4G, 4H, 4I, and 4J, the expression profiles of CiCaspase-9 and CiCaspase-3 were the same as we described previously [19]. To be specific, expression level of CiCaspase-9 and CiCaspase-3 in intestine was decreased at first upon GCRV stimulation and then began to up-regulate significantly around 4 or 5 days post stimulation ( Figure 4G and 4H).…”

“…After no abnormal symptom was observed, the grass carp was subjected to further study. Then the GCRV challenge experiments and samples collection were conducted as described before [19]. Briefly, viral suspension (4.24×10 6 TCID 50 /ml) was mixed an equal amount of commercial feed.…”

Germ Cells and Gonadal Development in a Teleost, Osteochilus vittatus (Valenciennes, 1842) Exposed to Potassium Dichromate

“…Even though Sivakumar et al (2014) research is based on mammals, Eimon and Ashkenazi (2010) approved of such proteins to be exist and have a same exact role in zebrafish. Wang et al (2016) also reported the presence of BAX in grass carp (Ctenopharyngodon idellus).…”