The Ciona myogenic regulatory factor functions as a typical MRF but possesses a novel N-terminus that is essential for activity.

Electroporation-based assays were used to test whether the myogenic regulatory factor (MRF) of Ciona intestinalis (CiMRF) interferes with endogenous developmental programs, and to evaluate the importance of its unusual N-terminus for muscle development. We found that CiMRF suppresses both notochord and endoderm development when it is expressed in these tissues by a mechanism that may involve activation of muscle-specific microRNAs. Because these results add to a large body of evidence demonstrating the exceptionally high degree of functional conservation among MRFs, we were surprised to discover that non-ascidian MRFs were not myogenic in Ciona unless they formed part of a chimeric protein containing the CiMRF N-terminus. Equally surprising, we found that despite their widely differing primary sequences, the N-termini of MRFs of other ascidian species could form chimeric MRFs that were also myogenic in Ciona. This domain did not rescue the activity of a Brachyury protein whose transcriptional activation domain had been deleted, and so does not appear to constitute such a domain. Our results indicate that ascidians have previously unrecognized and potentially novel requirements for MRF-directed myogenesis. Moreover, they provide the first example of a domain that is essential to the core function of an important family of gene regulatory proteins, one that, to date, has been found in only a single branch of the family.