Bombyx E75 isoforms display stage- and tissue-specific responses to 20-hydroxyecdysone.

Kang Li,Enen Guo,Xiaojuan Deng,Sheng Li,Ling Tian,Yang Cao,Qingrong Li,Muktadir S Hossain

doi:10.1038/srep12114

Abstract

Resulted from alternative splicing of the 5′ exons, the nuclear receptor gene E75 in the silkworm, Bombyx mori, processes three mRNA isoforms, BmE75A, BmE75B and BmE75C. From the early 5th larval instar to the prepupal stages, BmE75A mRNA and protein levels in the prothoracic glands display developmental profiles similar to ecdysteroid titer. In the fat body, mRNA levels but not protein levels of all three BmE75 isoforms correlate with ecdysteroid titer; moreover, proteins of all three BmE75 isoforms disappear at the prepupal stages, and a modified BmE75 protein with smaller molecular weight and cytoplasm localization occurs. At the early 5th larval instar stage, treatment of the prothoracic glands and fat body with 20-hydroxyecdysone (20E) and/or cycloheximide (CHX) revealed that BmE75A is 20E primary-responsive at both mRNA and protein levels, while BmE75B and BmE75C exhibit various responses to 20E. At the early wandering stage, RNAi-mediated reduction of gene expression of the 20E nuclear receptor complex, EcR-USP, significantly decreased mRNA and protein levels of all three BmE75 isoforms in both tissues. In conclusion, BmE75 isoforms display stage- and tissue-specific responses to 20E at both mRNA and protein levels; moreover, they are regulated by other unknown factors at the protein level.

Highlights

In Drosophila, there are four DmE75 mRNA isoforms, DmE75A, DmE75B, DmE75C and DmE75D, which originate from a single copy gene through differential promoter usage and alternative splicing of www.nature.com/scientificreports/
The AF-1 of BmE75A, BmE75B and BmE75C varies in sequence and length, implying that they recruit different co-activators or co-repressors onto target gene promoters to regulate gene expression in a ligand-independent manner[25]
The results show that, in the fat body, mRNA levels of all three BmE75 isoforms correlate with ecdysteroid titer

Summary

Introduction

In Drosophila, there are four DmE75 mRNA isoforms, DmE75A, DmE75B, DmE75C and DmE75D, which originate from a single copy gene through differential promoter usage and alternative splicing of www.nature.com/scientificreports/. Lose-of-function of DmE75B has no effects on DmHR3-induced DmβFtz-F1 expression and developmental transition[16] This could be because that DmE75A and DmE75B interact with DmHR3 to inhibit DmβFtz-F1 expression; and the function of DmE75A and DmE75B to inhibit the transcriptional activity of DmHR3 can be reversed by NO8,9. Similar to the findings in Drosophila, 20E rapidly and abundantly induces BmE75A expression in the ovary, while its induction of BmE75C expression is slow and weak. BmE75A expression proceeds with BmE75C expression during ovary development[22] Both BmE75A and BmE75C interact with BmHR3 and repress the transcriptional activity of BmHR3 to induce retinoic acid receptor-related receptor response element (RORE) linked target genes expression[23]. In addition to 20E, other unknown factors are involved in the regulation of BmE75 isoforms at the protein level

Methods

Results

Conclusion