Evidence from oyster suggests an ancient role for Pdx in regulating insulin gene expression in animals

Fei Xu,Xiao Liu,Peter W H Holland,Tatjana Sauka-Spengler,Daria Gavriouchkina,Ferdinand Marlétaz,Guofan Zhang

doi:10.1038/s41467-021-23216-7

Abstract

Hox and ParaHox genes encode transcription factors with similar expression patterns in divergent animals. The Pdx (Xlox) homeobox gene, for example, is expressed in a sharp spatial domain in the endodermal cell layer of the gut in chordates, echinoderms, annelids and molluscs. The significance of comparable gene expression patterns is unclear because it is not known if downstream transcriptional targets are also conserved. Here, we report evidence indicating that a classic transcriptional target of Pdx1 in vertebrates, the insulin gene, is a likely direct target of Pdx in Pacific oyster adults. We show that one insulin-related gene, cgILP, is co-expressed with cgPdx in oyster digestive tissue. Transcriptomic comparison suggests that this tissue plays a similar role to the vertebrate pancreas. Using ATAC-seq and ChIP, we identify an upstream regulatory element of the cgILP gene which shows binding interaction with cgPdx protein in oyster hepatopancreas and demonstrate, using a cell culture assay, that the oyster Pdx can act as a transcriptional activator through this site, possibly in synergy with NeuroD. These data argue that a classic homeodomain-target gene interaction dates back to the origin of Bilateria.

Highlights

Hox and ParaHox genes encode transcription factors with similar expression patterns in divergent animals
Two of the oyster genes consistently group with the Molluscan Insulin Peptide (MIP) group and are physically linked indicative of tandem duplication, as reported previously22. cgMILP7 is a likely orthologue of Drosophila[22], beetle and termite ILP7, a conclusion supported by their predicted chain structure (Supplementary Table 1)
Precise tree topology is dependent on alignment and sampling, and cgILP is placed less precisely in our analysis, we find that it groups with chordate insulin/insulin growth factor (IGF) genes, Bombyxins and spider genes (Fig. 1), supporting the close relationship with insulin[22]

Summary

Introduction

Hox and ParaHox genes encode transcription factors with similar expression patterns in divergent animals. Using ATAC-seq and ChIP, we identify an upstream regulatory element of the cgILP gene which shows binding interaction with cgPdx protein in oyster hepatopancreas and demonstrate, using a cell culture assay, that the oyster Pdx can act as a transcriptional activator through this site, possibly in synergy with NeuroD These data argue that a classic homeodomain-target gene interaction dates back to the origin of Bilateria. Chromatin immunoprecipitation suggests binding of Pdx protein, and luciferase assays in cell culture verifies the ability to activate gene expression through this non-coding site 1.8 kb upstream of the oyster cgILP gene

Methods

Results

Conclusion