Cloning and Expression of the Zebrafish Gene, Setd8, a Histone Methyl-tranferase Required for Cell Proliferation
Thompson, Shatia R.
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The epithelium is a basic type of animal tissue that provides a barrier to ensure homeostasis of internal environments in different external environments. Consequently zebrafish (Danio rerio) require an epithelium with limited permeability, the enveloping layer (EVL), early on to survive in a hypotonic external environment. The zebrafish EVL expresses distinct keratin genes and forms the external layer of the epidermis surrounding the embryo. It is hypothesize that the mechanism surrounding regulating epidermal differentiation is conserved between zebrafish and mammals, for this reason zebrafish have emerged as an experimental model. The poky/Ikk1 mutant has a defect in EVL differentiation causing the EVL to be hyper proliferative, undifferentiated and ineffective in forming a permeability barrier. Expression of ikk1 is lost in ~ 30% of human squamous cell carcinomas, indicating its continuing role of in maintaining epidermal homeostasis. Continued investigation of the zebrafish EVL will lead to understanding the mechanism for epithelial cell cycle regulation and differentiation. From an mRNA microarray experiment a histone methyl transferase, setd8, which is upregulated in poky mutants was identified. In this study, the setd8 gene was amplified out of poky cDNAs to determine periods of expression. The gene was broadly expressed in all stages of embryogenesis suggesting its importance in cell cycle progression and differentiation. In order to observe localization of expression an in situ hybridization was performed, yielding insignificant results. Prior to in situ hybridization the amplified gene setd8 sequence from poky cDNA was sequenced resulting in the right clone in comparison to hypothesized sequence. Sequence alignment with the human setd8 gene; indicate conservation of the setd8 gene between zebrafish and mammals.