The transcription factors Tbx6-r.b and Mrf form a positive feedback loop that is involved in regulating muscle cell specification in ascidians. In this new study, Oda and Satou investigate the role of the Tbx6-r.b/Mrf loop in the muscle cells of ascidians from the embryonic to larval stages. The authors identified 101 muscle-specific genes using publicly available single-cell transcriptomic data, and found that all muscle-specific genes are regulated under the control of Tbx6-r.b and Mrf. Through characterising the temporal expression profiles of muscle-specific genes and conducting morpholino knockdown experiments, the authors found that a group of transcription factor genes (including Snail, Meox and Tcf15-r) is under the control of the Tbx6-r.b/Mrf regulatory loop. These transcription factors together regulate the temporal expression patterns of muscle-specific genes. Specifically, Snail regulates muscle-specific genes in earlier stages, while Meox and Tcf15-r control gene expressions in late embryonic to larval stages. Adding Snail binding sites to a regulatory region of a gene that normally turns on later is sufficient to shift its onset of expression earlier. Overall, the findings suggest that Tbx6-r.b and Mrf form a master regulatory loop that activates all muscle-specific genes at different times, by coordinating the binding of upstream factors to cis-regulatory regions of downstream genes.