Expression and functional analysis of apoptosis-related gene BmICAD in silkworm, Bombyx mori
The ICAD gene plays an important role in the growth and development processes in insects. We conducted a molecular cloning and functional analysis to study a specific silkworm gene BmICAD related to apoptosis. The BmICAD gene was obtained from the fifth instar larvae of the silkworm by RTPCR and over-expressed in Escherichia coli as His-tagged fusion proteins. Subcellular localization of the protein indicated that BmICAD was found in the cytoplasm near the nucleus. RNAi assay indicated that the apoptosis rate of Bm5 cells increased markedly. A His pull-down assay was used to investigate proteins that bind to rBmICAD. Two proteins, DNA supercoiling factor (SCF) and secreted protein acidic and rich in cysteine (SPARC), were found. These results indicate that down regulation of BmICAD can increase the apoptosis rate of Bm5 cells, and that SCF and SPARC may have important roles by interacting with BmICAD during this process.
Key words: Bombyx mori, BmICAD, subcellular localization, RNAi, protein interaction.