Abstract: 【Objective】Yki, a transcriptional coactivator, is phosphorylated and activated by upstream kinases cascade of the Hippo signaling pathway, and then regulates the downstream genes expression to control organ size, proliferation and apoptosis of cell. As a model of Lepidopteran insect, little is known about the Hippo signaling pathway involved in the silkworm (Bombyx mori). The objective of this study is to identify B. mori BmYki-1 and analyze its sequence information and express characteristics, laying the foundation for further study of Hippo signaling pathways in midgut, which will help us better understand Hippo signaling pathways in intestinal regulatory mechanism of steady maintaining and tissue damage repairing.【Method】Based on mammals and drosophila Yki protein sequence, B. mori homologous Yki gene sequences was identified using NCBI databases and silkworm genome database. B. mori Dazao strain being research material, full-length sequence of two Yki spliced variants were cloned by PCR and RACE. According to bioinformatics methods, gene sequences and protein structure characteristics of the two spliced variants were analyzed. Here, function of BmYki-1 in B. mori intestinal tissue was focused. The spatial-temporal expression profile of BmYki-1 was investigated by semi-quantitative RT-PCR. BmYki-1 was constructed into a transit expression vector, and transfected into B. mori embryonic cell line to analyze the subcellular location of BmYki-1. qRT-PCR was utilized to detect the expression of BmYki-1 in B. mori intestinal after treated with bleomycin.【Result】There are two alternatively spliced forms of BmYki, namely BmYki-1 and BmYki-2, whose Gene ID are BGIBMGA0082-TA and BGIBMGA0081-TA in silkworm genome database. The ORF of the two spliceosomes consist of 1 329 and 1 227 bp, encoding 442 and 408 amino acids, with 48 and 44.1 kD protein molecular weight and 5.18 and 5.50 isoelectric point, respectively. In addition, both spliceosomes have two WW domains from 139th to 171th and 220th to 252th amino acids. A multi species phylogenetic tree demonstrated that BmYki-1 was most closely related to Amyelois transitella, which is Lepidoptera insect like B. mori, and was comparatively closely related to Coleoptera and Hymenoptera insects. However, BmYki-1 had a distant relationship with that of Diptera insect, and furthest with vertebrates. Expression profile showed that BmYki-1 was specifically expressed in the midgut and silk gland, and the expression was significantly up-regulated from 2nd molting stage in the midgut and reached the top at early-to-mid stage of 5th instar. Overexpressed BmYki-1 in BmE cells was mainly distributed in the nucleus. BmYki-1 was significantly up-regulated in the midgut treated with bleomycin compared to the control.【Conclusion】The BmYki-1 cDNA was cloned and its expression patterns in B. mori were analyzed. The subcellular location of BmYki-1 was investigated and mainly located in nucleus. BmYki-1 may play a critical role in the development and repair of damaged midgut tissue of B. mori.

Key words: silkworm (Bombyx mori), BmYki-1, bioinformatics analysis, spatial-temporal expression profile, subcellular localization