Crosstalk between vrille transcripts, proteins and regulatory elements controlling circadian rhythms and development in Drosophila

The vrille (vri) gene encodes a bZIP transcriptional repressor that is required for Drosophila patterning and cell proliferation during development as well as circadian behavior in adults. Three alternate first exons produce five different vri transcripts, where first exons 1a and 1b initiate two transcripts during development that produce the 610aa short VRI isoform and first exon 1c initiates three transcripts in adults that produce the 729aa long VRI isoform. To test whether long VRI is necessary for circadian clock function, we generated a mutant (vriΔ679) that eliminates transcripts encoding long VRI. The vriΔ679 mutant is viable, confirming that developmental functions of vri are driven by transcripts from exons 1a/1b, yet behavioral rhythms persist in vriΔ679 flies, demonstrating that vri transcripts encoding short VRI are sufficient for clock output. E-box regulatory elements upstream of exon 1c that drive rhythmic transcription in adults are also required for development, implying that these E-boxes also drive vri transcription from exons 1a/1b. Surprisingly, there is almost no long VRI present in adults even though the vast majority of transcripts encode long VRI, apparently because the ATG start codon for long VRI has a poor Kozak sequence context. Consequently, short VRI is the predominant isoform in adults and is sufficient for clock output. We conclude that vri-ADF transcripts driven by E-boxes upstream of exon 1c primarily control circadian rhythms by producing short VRI , whereas vri-E transcripts that are also regulated by E-boxes upstream of exon 1c mediate developmental patterning/cell proliferation through short VRI. RNA-seq: mRNA profiles of heads from Drosophila wild type (WT) and vriΔ679 mutant at two different timepoints, ZT 02 and ZT 14