MEF2D regulates the expression of mesoderm genes.
(A) qPCR analysis of embryos injected with MEF2D AMO or mismatch AMO. 5 embryos on each group were injected and RNA was extracted at stage 9. qPCR was performed as described in “materials and methods”. Expression levels of each gene were arbitrarily set to a value of 1 in the mismatch AMO injected embryos. The values for each gene were standardized accordingly. Data are presented as means ± SE of two independent experiments with duplicates(B) qPCR was performed on stage 10.5 embryos treated as is described in A. (C) Stage 10.5 embryos were analyzed by ISH using antisense probes to brachyury, goosecoid and chordin (left to right). (D) Control and MEF2D AMO-injected embryos were analyzed at stage 14 by RT-PCR (left) and at stage 16 by ISH with antisense probe to myod. (E) Transversal sections of stage 24 control and MEF2D AMO-injected embryos analyzed by immunohistochemistry. The antibody used was anti Myosin heavy chain (MF20) and samples were counterstained with hematoxylin. (F) Co-injection of MEF2D-Flag mRNA with MEF2D-AMO restores the expression of mesoderm and organizer genes. MEF2D AMO was injected alone or together with Mef2D-Flag mRNA into one cell embryos. Embryos were analyzed by RT-PCR (left) and by ISH with antisense probe to goosecoid (right) at stage 10.25.