her15, a novel gene with oscillating mRNA expression domains and its potential role in zebrafish somitogenesis

Somitogenesis is the key developmental process which lays down the framework for an organised body plan in vertebrates and cephalochordates. Somitogenesis divides the body axis into transient segmental structures called somites, which later give rise to muscles and vertebrae of the trunk and tail. Somites are generated from the unsegmented presomitic mesoderm (PSM) by an intricate process of prepatterning. Prepatterning is driven by a segmentation clock referred to as the presomitic mesoderm oscillator. This oscillator consists of certain gene members with oscillating mRNA expression compartments that sweep like a wave from the posterior to the anterior end of the embryonic PSM. The Delta-Notch pathway and various genes belonging to the hairy-(h) and Enhancer of split � [E(spl)] related family, are the core conserved components of this oscillator. h/E(spl) genes in zebrafish are commonly referred to as her genes. her1 and her7, play very important roles in the regulation of somitogenesis. The open question is how many her genes are core components of the zebrafish presomitic mesoderm oscillator and how do they interact with one another? To answer this, an in situ screen for h/E(spl) genes in zebrafish was conducted by Sieger et al., (2004). Three new her genes with oscillating mRNA expression domains were identified and one of them is her15, which has been further characterized in this PhD thesis. her15 mRNA is expressed as a distinct oscillatory posterior PSM domain which shows three primary phases, namely broad, intermediate and dot-like. Comparable to other her genes, her15 also showed stripes in the anterior PSM but unlike others, these stripes were found to be expressed at double segmental distance. Additionally, the her15 stripe was found to label the posterior border of the last somite. Morpholino gene knock down studies showed that the oscillating expression of her15 is partly dependent on her7 regulation, but independent of her1. Oscillating her15 mRNA signals in the posterior PSM displays fluctuations with respect to left and right halves of the embryonic PSM, suggestive of the autonomy of both halves of the embryo in generating the signal. Misexpression studies suggest a prospective role for her15 in the regulation of somite border formation and oscillatory gene expression in zebrafish PSM. Morpholinos against her15 did not result in morphological border disruption, or in changes in mRNA expression of genes of the Delta-Notch pathway. ZfChp, the second candidate gene which has been analyzed in the present thesis, came out of a screen of the NIH cDNA in situ expression database. ZfChp exhibits dynamic stripes of mRNA expression in the intermediate PSM region, a dynamic zone where tail bud mesenchymal cells undergo transition to epithelial state (mesenchymal to epithelial transition-MET), thus giving rise to somites. It the molecular signature of a Rho family GTPase with respect to conserved Rho GTPase domains. The Delta-Notch signaling pathway positively regulates the dynamic stripes of ZfChp in the intermediate PSM region. This provides the first molecular evidence supporting a link between prepatterning of the PSM and MET in zebrafish embryos.