Identification and Characterization of a Novel Microvitellogenin from the Chinese Oak Silkworm Antheraea pernyi.

Yanqun Liu,Qun Li,Junfang Su,Xixi Zheng,Shenglin Shi,Hongfang Ma,Miaomiao Chen,Li Qin,Kenneth Söderhäll

doi:10.1371/journal.pone.0131751

Abstract

Microvitellogenin (mVg) is a relatively small vitellogenic protein only characterized in the eggs of the lepidopteran insects Manduca sexta and Bombyx mori. In the present study, we report a novel mVg (ApmVg) isolated from the Chinese oak silkworm Antheraea pernyi. The obtained ApmVg cDNA sequence contains an open reading frame of 783 bp encoding a protein of 260 amino acids with a predicted molecular weight of 29.96 kDa. This gene does not contain introns. Structural analysis revealed that this protein shares putative conserved domains with the lepidopteran low-molecular weight lipoprotein, which belongs to the lipoprotein_11 superfamily. The protein sequence of ApmVg exhibits 48% sequence identity with mVg from M. sexta and 40–47% sequence identity with the 30K lipoproteins from B. mori. Phylogenetic analysis suggests that ApmVg is a novel member of the lepidopteran low-molecular weight lipoproteins. Transcriptional analysis indicated that ApmVg mRNA is mainly expressed in the fat body (both female and male) during post-diapause development of the pupal stage, and it was also detected in ovaries and spermaries in smaller amounts. RT-PCR and Western blot analyses revealed that ApmVg is synthesized by the fat body and secreted into hemolymph and ultimately accumulates in eggs. The ApmVg transcript can be detected in the fat bodies of female pupae four days after treatment with 20-hydroxyecdysone and shows an expression pattern distinct from that of vitellogenin (Vg), which is detectable throughout diapausing and in post-diapause development. ApmVg decreased dramatically during embryonic development. These results represent the first study of mVg outside M. sexta and B. mori and provide insight into the physiological role and evolution of mVgs.

Highlights

In animals, yolk proteins are essential to ensure a sufficient supply of nutrients for the developing embryo [1]
The obtained 928 bp cDNA sequence contains a 5’-untranslated region (UTR) of 40 bp, a 3’ UTR of 105 bp with a polyadenylation signal sequence AATAAA at position 824 and a poly (A) tail, and an open reading frame (ORF) of 783 bp encoding a polypeptide of 260 amino acids (S1 Fig)
Protein sequence comparison revealed that the characterized protein in this study shared 48% identity with the known mVg isolated from M. sexta. mRNA expression and Western blot analysis demonstrated that mVg is synthesized by the fat body, secreted into hemolymph, and accumulates in the eggs of A. pernyi, as previously described for M. sexta mVg [9]

Summary

Introduction

Yolk proteins are essential to ensure a sufficient supply of nutrients for the developing embryo [1]. Vgs are large molecules (approximately 200 kDa) synthesized by the fat body, secreted into the hemolymph, and transported to the developing oocytes. MVg is a female-specific yolk protein in moths, and mVg mRNA is only present in the adult female fat body [7]. This protein accumulates in the eggs, presumably via an endocytic process, but mVg does not use the same endocytic receptor as that of Vg [8]. We report a novel mVg from the Chinese oak silkworm Antheraea pernyi (Lepidoptera: Saturniidae). Our data could help delineate the physiological functions of these related proteins

Materials and Methods

Results

Discussion and Conclusions