Bioinformatic meta-analysis of transcriptomics of developing Drosophila muscles identifies temporal regulatory transcription factors including a Notch effector

Abstract

The intricate mechanism of gene regulation coordinates the precise control of when, where, and to what extent genes are activated or repressed, directing the complex processes that govern cellular functions and development. Dysregulation of gene expression can lead to diseases such as autoimmune disorders, cancer, and neurodegeneration. Transcriptional regulation, especially involving transcription factors (TFs), plays a major role in controlling gene expression. This study focuses on identifying gene regulatory mechanisms that generate distinct gene expression patterns during Drosophila muscle development. Utilising a bioinformatics approach, we analysed the developmental time-point-specific transcriptomics resource generated by Spletter et al., which includes mRNA sequencing data at eight stages of indirect flight muscle (IFM) development. They had identified 40 distinct genome-wide clusters representing various temporal expression dynamics using 'soft' clustering. Promoter sequences of genes in these clusters were analysed to predict novel motifs that act as TF binding sites. Comparative analysis with known motifs revealed significant overlaps, indicating shared transcriptional regulation. The physiological relevance of predicted TFs was confirmed by cross-referencing with experimental ChIP-seq data. We focused on Cluster 36, characterised by a unique bimodal temporal expression profile, and identified candidate genes, Rbfox1 and zfh1, for further study. Ectopic overexpression experiments revealed that the TF Enhancer of split m8 helix-loop-helix [E(spl)m8-HLH], part of the Notch signalling pathway, acts as a transcriptional repressor for Rbfox1 and zfh1. Our findings highlight the complexity of transcriptional regulation during myogenesis, and identify key TFs that could be targeted for further research in muscle development and related disorders.