Identification and subcellular localization analysis of CCTα in microsporidian Nosema bombycis.

Abstract

CCT is a chaperonin which is widely present in eukaryotic cells and mainly involves in the folding and assembly of cytoskeletal proteins β-tubulin and actin. The alpha subunit of CCT(CCTα) plays a pivotal role in the folding and assembly of cytoskeletal protein(s) as an individuals or complexes. In this study, we report cloning, characterization and expression of the CCTα of Nosema bombycis (NbCCTα) for the first time. The NbCCTα gene contains a complete ORF of 1629bp in length that encodes a 542-amino acid polypeptide. The NbCCTα is 59.662kDa molecular weight in size with an isoelectric point (pI) of 5.81, no signal peptide or transmembrane domain. The IFA results showed that the NbCCTα was co-localized with actin and β-tubulin in the cytoplasm, nucleus, nuclear membrane and plasma membrane of N. bombycis in the process of proliferation. qPCR analysis showed that the relative expression level of NbCCTα increased from 24h to 96h post-infection (hp.i) of N. bombycis, and reached the highest at 96hp.i. The relative expression level of NbCCTα gene after RNAi was restrained at a low level from 48hp.i to 96hp.i. Knockdown of NbCCTα gene down-regulated the expression of Nbβ-tubulin and Nbactin genes. These results imply that NbCCTα may play an important role in the lifecycle of N. bombycis.