Cloning and characterization of homologue of mitotic arrest deficient 2 from Pyropia haitanensis and its expression analysis during parthenogenesis

To investigate the role and molecular mechanism of diploidization of cells during the parthenogenetic development of female gametophyte in Pyropia haitanensis, a cDNA sequence of homologue of mad2 was isolated and its expression profile at different developmental stages of parthenogenesis was characterized. In this study, the cDNA of Ph-mad2 spans 672 bp that encodes a protein of 223 amino acids (aa) with a predicted molecular weight of 23.8 ku. The deduced protein sequence of PhMAD2 has a typical HORMA domain and conservative serine and threonine phosphorylation sites. Phylogenetic analysis revealed that evolutionary status of P. haitanensis was extremely related to the fungi from Phycomycetes, such as Saprolegnia diclina and S. parasitica. Quantitative real-time PCR (qRT-PCR) analysis showed that the expressions of Ph-mad2 at reproductive cell development stage (RDS) and parthenospore development stage (PDS) with diploid carpogonium-like cells and carpospore-like cells were significantly down-regulated compared with that at vegetative cell proliferation stage (VPS) with haploid vegetative cells, while its expression profiles were up-regulated in the later stages of parthenosporophyte formation stage (PFS) and parthenosporophyte growth stage (PGS), in which the diploid carpospore-like germlings and parthenosporophytes underwent normal mitosis. These results suggested that Ph-mad2 reduction potentially prevented the formation of spindle assembly checkpoint (SAC) at mitotic cells during the early stages of parthenogenesis, and triggered their development of diploidization. These results provide important information for revealing the mechanism of spontaneous chromosome doubling in the parthenogenesis of P. haitanensis.