March 1, 2002;
eFGF is required for activation of XmyoD expression in the myogenic cell lineage of Xenopus laevis.
This paper addresses the molecular mechanisms that regulate the transcriptional activation of the myogenic regulatory factor XmyoD
in the skeletal muscle
lineage of Xenopus laevis. Using antisense morpholino oligonucleotide-mediated inhibition, we show that the signalling molecule embryonic fibroblast
growth factor (eFGF
), which is the amphibian homologue of FGF4
, is necessary for the initial activation of XmyoD
transcription in myogenic cells. We demonstrate that eFGF
can activate the expression of XmyoD
in the absence of protein synthesis, indicating that this regulation is direct. Our data suggest that regulation of XmyoD
expression may involve a labile transcriptional repressor. In addition, we show that eFGF
is itself an immediate early response to activin, a molecule that mimics the endogenous mesoderm
-inducing signal. We propose a model for the regulation of XmyoD
within the early mesoderm
, and discuss the relevance that these findings have for the understanding of myogenic specification in higher vertebrates.
[+] show captions
Fig. 1. The normal expression patterns of eFGF and XmyoD showing co-expression in the early mesoderm. Whole-mount in situ hybridisation showing expression of (A) eFGF at stage 10, (B) XmyoD at stage 10 and (C) XmyoD at stage 10 (+). Expression of XmyoD across the dorsal midline is rapidly excluded as Spemann’s organiser signalling is established.
Fig. 5. eFM inhibits XmyoD expression in gastrula but not neurula embryos. (A) (i) RNA was extracted at stages 11 and 14 from whole embryos, embryos injected with 80 ng, and embryos injected with 40, 60 or 80 ng of eFM. Total RNA (10 μg) was assayed by RNAase protection for expression of the mesodermal markers XmyoD and Xsna and for the loading control ODC; (ii) RNA was extracted from whole embryos at stage 13, embryos injected with 80 ng of eFM, embryos injected with 200 pg of ssbFGF, and embryos injected with 80 ng of eFM and 200 pg of ssbFGF. Total RNA (10 μg) was analysed by RNAase protection for expression of the marker genes XmyoD, Xbra and the loading control ODC. (B) Embryos injected unilaterally on the left hand side with 40 ng of eFM were assayed at (i) gastrula stage 10.5 and (ii) neurula stage 14 for XmyoD gene expression by in situ hybridisation. (iii) Gastrula stage 10.5 embryos injected unilaterally on the left-hand side with 40 ng of eFM were also assayed by in situ hybridisation for expression of the mesodermal marker Xsna.