Identification, characterization and functional analysis of anti-apoptotic protein BCL-2-like gene from pufferfish, Takifugu obscurus, responding to bacterial challenge

Abstract

Apoptosis plays a crucial role in many biological processes, including development, cellular homeostasis and immune responses. The BCL-2 family is a key regulator of the mitochondrial response to apoptotic signals in the intrinsic pathway. In this study, we identified and characterized the cDNA and expression pattern of pufferfish BCL-2 (PfBCL-2). The full-length cDNA of PfBCL-2 was 1412 bp with an open reading frame of 657 bp encoding a putative protein of 219 amino acids (Accession no: KP898414). The calculated molecular mass of the PfBCL-2 was 24.2 kDa with a predicted isoelectric point of 5.27. The deduced PfBCL-2 protein exhibited four highly conserved BCL-2 homology domains, suggesting that PfBCL-2 may play a similar role in the apoptotic-signaling pathway as in other species. Real-time PCR results showed that PfBCL-2 transcript was expressed in a wide range of tissues but exhibited the greatest level of expression in blood. Transcriptional responses of PfBCL-2 exhibited different spatial and temporal expression profiles in liver and blood after bacterial infection. PfBcl-2 transcript was significantly up-regulated in liver at 6, 12, 24 and 48 h (with maximum induction at 48 h) and was up-regulated in blood at 3, 6, 12 and 24 h (with maximum induction at 12 h). Meanwhile, recombinant PfBCL-2 fused with His6 tag was efficiently expressed in Escherichia coli BL21 (DE3) and purified using Ni-nitrilotriacetic acid resin. Western blot analysis indicated that its protein level appeared to be elevated during the initial bacterial infection. These results suggest that PfBCL-2 plays important roles in immune responses against bacteria challenge.

Functional characterization of TNF‐α in pufferfish (Takifugu obscurus) in immune response and apoptosis against Aeromonas hydrophila

“…The pufferfish ( Takifugu obscurus ) is anadromous and distribute in East China Sea, Yellow Sea and Sea of Japan with considerable economic value (Cheng et al., 2015). In recent years, bacterial diseases have brought a negative impact on the aquaculture industry, including the gram‐negative bacterium Aeromonas hydrophila , the main pathogen of pufferfish (Fu et al., 2020).…”

Molecular cloning, characterization and expression analysis of major histocompatibility complex class I alpha gene of pufferfish (Takifugu obscurus)

“…However, the exact mechanism by which A. hydrophila modulates the host immune response in fish to their advantage and the specific factors that contribute to the pathogenesis are poorly understood. Takifugu obscurus , commonly known as pufferfish, is a commercially important fish along the coastal regions of China, Japan and Korea (Yang & Chen, 2003), and A. hydrophila is the main pathogen of T. obscures (Cheng et al., 2015). In this study, a comparative analysis of cDNA and protein sequences of pufferfish MHC Ⅰα named PfMHC Ⅰα was reported.…”

Molecular characterization and expression analysis of B-cell lymphoma-2 in Trachinotus ovatus and its role in apoptotic process

“…To date, several Bcl-2 homologous sequences have been cloned and characterized in teleosts, such as Danio rerio, Channa striatus, Takifugu obscurus, Ictalurus punctatus, and Epinephelus coioides (2,4,(32)(33)(34). The anti-apoptotic function of fish Bcl-2 seems to be conserved compared with that of mammalian Bcl-2.…”