Six1 regulates proliferation of Pax7+ muscle progenitors in zebrafish

In the embryonic zebrafish, skeletal muscle fibres are formed from muscle progenitors in the paraxial mesoderm. The embryonic myotome is mostly constituted of fast twitch specific fibres, which are formed from a fast specific progenitor cell pool. The most lateral fraction of the fast domain in the myotome of zebrafish embryos derive from the Pax7 positive dermomyotome-like cells. In this study we show that two genes, belonging to the sine oculus class 1 (six1) genes (six1a and six1b) both are essential for the regulation of Pax7(+) cell proliferation and consequently in their differentiation during the establishment of the zebrafish dermomyotome. Both in six1a and in six1b morphant embryos, Pax7(+) cells are initially formed, but fail to proliferate as detected by the reduced proliferation marker phosphohistone3 and brdU incorporation. In congruence, overexpression of six1a or six1b leads to increased Pax7(+) cell number and reduced or alternatively delayed fibre cell differentiation. BMP signalling has previously been suggested to inhibit differentiation of Pax7(+) cells in the dermomyotome. Here we show that the remaining Pax7(+) cells in six1a and six1b morphant embryos also have significantly reduced pSmad1/5/8 levels and propose that this leads to a reduced proliferative activity, which may result in a premature differentiation of Pax7(+) cells in the zebrafish dermomyotome. In summary, we show a mechanism for Six1a and Six1b in establishing the Pax7(+) cell derived part of the fast muscle and suggest new important roles for Six1 in the regulation of the Pax7(+) muscle cell population through pSmad1/5/8 signalling.