title: Bmo-miR-2758 Targets BmFMBP-1 (Lepidoptera: Bombycidae) and Suppresses Its Expression in BmN Cells.
abstract: MicroRNAs (miRNAs) are an abundant family of endogenous noncoding small RNA molecules. They play crucial roles on regulation of life processes both in plants and animals. Fibroin modulator binding protein-1 (FMBP-1) is a silk gland transcription factor of Bombyx mori, which is considered as a trans-activator of fibroin genes. And bioinformatics prediction showed that at the 3' untranslated region (3' UTR) of BmFMBP-1 there were binding sites for three bmo-miRNAs, bmo-miR-2b*, bmo-miR-305, and bmo-miR-2758, separately. In order to validate whether these bmo-miRNAs involved in the regulation of BmFMBP-1 expression, the expression levels of three bmo-miRNAs and BmFMBP-1 in the middle silk gland (MSG) and posterior silk gland (PSG) during the fourth- and fifth-larval stages of B. mori were measured by semi-quantitative reverse transcription polymerase chain reaction. The results revealed that the expression level of bmo-miR-2758 was the highest in the three, and it expressed higher in the PSG than in the MSG with a similar expression pattern as BmFMBP-1, implying that bmo-miR-2758 may involved in regulation of BmFMBP-1. To validate the regulation function of bmo-miR-2758 on BmFMBP-1, recombinant plasmids pcDNA3 [ie1-egfp-pri-bmo-miR-2758-SV40] and pGL3 [A3-luc-FMBP-1 3' UTR-SV40] were constructed and co-transfected in BmN cells. The dual-luciferase reporter assay system was used for assay of transient expression. The results showed that the expression of the luciferase reporter was significantly decreased when pGL3 [A3-luc-FMBP-1 3' UTR-SV40] co-transfected with pcDNA3 [ie1-egfp-pri-bmo-miR-2758-SV40] (P <  .01). Furthermore, when the artificial antisense RNA of bmo-miR-2758 (inhibitor) was added to the above co-transfection, the expression of the luciferase reporter was recovered significantly (P < 0.01). These results suggest that bmo-miR-2758 represses the expression of BmFMBP-1 in vitro.
authors.affiliation: Jiangsu Key Laboratory of Sericultural Biology and Biotechnology, School of Biotechnology, Jiangsu University of Science and Technology, Zhenjiang, China; Key Laboratory of Silkworm and Mulberry Genetic Improvement, Ministry of Agriculture, Sericultural Research Institute, Chinese Academy of Agricultural Sciences, Zhenjiang, China.
authors.forename: Xin
authors.initials: X
authors.lastname: Wang
authors.affiliation: Jiangsu Key Laboratory of Sericultural Biology and Biotechnology, School of Biotechnology, Jiangsu University of Science and Technology, Zhenjiang, China; Key Laboratory of Silkworm and Mulberry Genetic Improvement, Ministry of Agriculture, Sericultural Research Institute, Chinese Academy of Agricultural Sciences, Zhenjiang, China.
authors.forename: Shunming
authors.initials: S
authors.lastname: Tang
authors.affiliation: Jiangsu Key Laboratory of Sericultural Biology and Biotechnology, School of Biotechnology, Jiangsu University of Science and Technology, Zhenjiang, China;
authors.forename: Fei
authors.initials: F
authors.lastname: Song
authors.affiliation: Jiangsu Key Laboratory of Sericultural Biology and Biotechnology, School of Biotechnology, Jiangsu University of Science and Technology, Zhenjiang, China;
authors.forename: Chen
authors.initials: C
authors.lastname: Chen
authors.affiliation: Jiangsu Key Laboratory of Sericultural Biology and Biotechnology, School of Biotechnology, Jiangsu University of Science and Technology, Zhenjiang, China; Key Laboratory of Silkworm and Mulberry Genetic Improvement, Ministry of Agriculture, Sericultural Research Institute, Chinese Academy of Agricultural Sciences, Zhenjiang, China.
authors.forename: Xijie
authors.initials: X
authors.lastname: Guo
authors.affiliation: Jiangsu Key Laboratory of Sericultural Biology and Biotechnology, School of Biotechnology, Jiangsu University of Science and Technology, Zhenjiang, China; Key Laboratory of Silkworm and Mulberry Genetic Improvement, Ministry of Agriculture, Sericultural Research Institute, Chinese Academy of Agricultural Sciences, Zhenjiang, China shenxjsri@163.com.
authors.forename: Xingjia
authors.initials: X
authors.lastname: Shen
mesh_terms: D000818:Animals; D012831:Bombyx / Q000235:genetics* / Q000378:metabolism; D005786:Gene Expression Regulation / Q000235:genetics / Q000502:physiology; D019476:Insect Proteins / Q000378:metabolism* / Q000502:physiology; D035683:MicroRNAs / Q000378:metabolism* / Q000502:physiology; D016133:Polymerase Chain Reaction; D014157:Transcription Factors / Q000378:metabolism / Q000502:physiology
chemical_list: D019476:Insect Proteins; D035683:MicroRNAs; D014157:Transcription Factors
keywords: BmFMBP-1; Bombyx mori; Functional identification; Silk gland transcription factor; microRNA