Mutation of a Cuticle Protein Gene, BmCPG10, Is Responsible for Silkworm Non-Moulting in the 2nd Instar Mutant.
In the silkworm, metamorphosis and moulting are regulated by ecdysone hormone and juvenile hormone. The subject in the present study is a silkworm mutant that does not moult in the 2nd instar (nm2). Genetic analysis indicated that the nm2 mutation is controlled by a recessive gene and is homozygous lethal. Based on positional cloning, nm2 was located in a region approximately 275 kb on the 5th linkage group by eleven SSR polymorphism markers. In this specific range, according to the transcriptional expression of thirteen genes and cloning, the relative expression level of the BmCPG10 gene that encodes a cuticle protein was lower than the expression level of the wild-type gene. Moreover, this gene's structure differs from that of the wild-type gene: there is a deletion of 217 bp in its open reading frame, which resulted in a change in the protein it encoded. The BmCPG10 mRNA was detectable throughout silkworm development from the egg to the moth. This mRNA was low in the pre-moulting and moulting stages of each instar but was high in the gluttonous stage and in newly exuviated larvae. The BmCPG10 mRNA showed high expression levels in the epidermis, head and trachea, while the expression levels were low in the midgut, Malpighian tubule, prothoracic gland, haemolymph and ventral nerve cord. The ecdysone titre was determined by ELISA, and the results demonstrated that the ecdysone titre of nm2 larvae was lower than that of the wild-type larvae. The nm2 mutant could be rescued by feeding 20-hydroxyecdysone, cholesterol and 7-dehydrocholesterol (7dC), but the rescued nm2 only developed to the 4th instar and subsequently died. The moulting time of silkworms could be delayed by BmCPG10 RNAi. Thus, we speculated that the mutation of BmCPG10 was responsible for the silkworm mutant that did not moult in the 2nd instar.
School of Biotechnology, Jiangsu University of Science and Technology, Zhenjiang, 212018, China.|The Sericulture Research Institute, Chinese Academy of Agricultural Sciences, Zhenjiang, 212018, China.|Industrial Crops Institute, Hubei Academy of Agricultural Sciences, Wuhan, 430064, China.
Fan
F
Wu
School of Biotechnology, Jiangsu University of Science and Technology, Zhenjiang, 212018, China.|The Sericulture Research Institute, Chinese Academy of Agricultural Sciences, Zhenjiang, 212018, China.
Pingyang
P
Wang
School of Biotechnology, Jiangsu University of Science and Technology, Zhenjiang, 212018, China.|The Sericulture Research Institute, Chinese Academy of Agricultural Sciences, Zhenjiang, 212018, China.
Qiaoling
Q
Zhao
School of Biotechnology, Jiangsu University of Science and Technology, Zhenjiang, 212018, China.|The Sericulture Research Institute, Chinese Academy of Agricultural Sciences, Zhenjiang, 212018, China.
Lequn
L
Kang
School of Biotechnology, Jiangsu University of Science and Technology, Zhenjiang, 212018, China.|The Sericulture Research Institute, Chinese Academy of Agricultural Sciences, Zhenjiang, 212018, China.
Dingguo
D
Xia
School of Biotechnology, Jiangsu University of Science and Technology, Zhenjiang, 212018, China.|The Sericulture Research Institute, Chinese Academy of Agricultural Sciences, Zhenjiang, 212018, China.
Zhiyong
Z
Qiu
School of Biotechnology, Jiangsu University of Science and Technology, Zhenjiang, 212018, China.|The Sericulture Research Institute, Chinese Academy of Agricultural Sciences, Zhenjiang, 212018, China.
Shunming
S
Tang
School of Biotechnology, Jiangsu University of Science and Technology, Zhenjiang, 212018, China.|The Sericulture Research Institute, Chinese Academy of Agricultural Sciences, Zhenjiang, 212018, China.
Muwang
M
Li
School of Biotechnology, Jiangsu University of Science and Technology, Zhenjiang, 212018, China.|The Sericulture Research Institute, Chinese Academy of Agricultural Sciences, Zhenjiang, 212018, China.
Xingjia
X
Shen
School of Biotechnology, Jiangsu University of Science and Technology, Zhenjiang, 212018, China.|The Sericulture Research Institute, Chinese Academy of Agricultural Sciences, Zhenjiang, 212018, China.
Guozheng
G
Zhang
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D012831:Bombyx / Q000737:chemistry / Q000235:genetics* / Q000502:physiology*; D002784:Cholesterol / Q000378:metabolism; D005260:Female; D017344:Genes, Insect; D019476:Insect Proteins / Q000032:analysis / Q000235:genetics*; D008297:Male; D008969:Molecular Sequence Data; D018978:Molting; D009154:Mutation
D019476:Insect Proteins; D002784:Cholesterol