Myogenin mutants have reduced expression of fusogenic factors. a ISH on 20?hpf myogkg128/+ incross to investigate the expression of genes essential for vertebrate myocyte fusion. Expression of myomaker (mymk) and jam3b, but not of jam2a or kirrel3l, is reduced in myog mutant. Bar?=?50??m. Representative images n?=?10 mutants, n?=?38 sibs (mymk); n?=?13 mutants, n?=?37 sibs (jam2a); n?=?12 mutants, n?=?39 sibs (jam3b); n?=?7 mutants, n?=?13 sibs (kirrel3l). b qPCR analysis of RNA expression levels showing downregulation of mymk, myomixer/myomerger/minion (mymx) and jam3b at 20?hpf on myogkg125 mutants, whereas jam2a and kirrel3l remain unaltered compared to wt (myog+/+) sibs. Graphs show mean fold change?±?SEM of four independent experiments; paired t test. Symbol shapes denote wt and mutant samples from paired experiments. c Schematic of 5' genomic region of mymk reporting: 5?-UTR and coding sequence (grey and white boxes), position of E-box elements (red boxes), relative bp distance from 5'UTR start (+1), start codon (arrow, ATG). E-box 1 and E-box 2 sequences are shown in red text. d ChIP-qPCR assay using anti-Myog or mock showing significant Myogenin enrichment on mymk E-box 1 compared to negative controls from the gapdh promoter and a gene-free region on chromosome 14 containing an E-box (Chr14). Mean of percentage of input immunoprecipitated?±?SD of two independent experiments, ANOVA
|
