Abstract
BackgroundEstablishment of the left-right axis is important for positioning organs asymmetrically in the developing vertebrate-embryo. A number of factors like maternally deposited molecules have emerged essential in initiating the specification of the axis; the downstream events, however, are regulated by signal-transduction and gene-expression changes identifying which remains a crucial challenge. The EGF-CFC family member Cryptic, that functions as a co-receptor for some TGF-beta ligands, is developmentally expressed in higher mammals and mutations in the gene cause loss or change in left-right axis asymmetry. Despite the strong phenotype, no transcriptional-regulator of this gene is known till date.ResultsUsing promoter-analyses tools, we found strong evidence that the developmentally essential transcription factor Snail binds to the human Cryptic-promoter. We cloned the promoter-region of human Cryptic in a reporter gene and observed decreased Cryptic-promoter activation upon increasing Snail expression. Further, the expression of Cryptic is down-regulated upon exogenous Snail expression, validating the reporter assays and the previously identified role of Snail as a transcriptional repressor. Finally, we demonstrate using gel-shift assay that Snail in nuclear extract of PANC1 cells interacts with the promoter-construct bearing putative Snail binding sites and confirm this finding using chromatin immunoprecipitation assay.ConclusionsSnail represses the expression of human Cryptic and therefore, might affect the signaling via Nodal that has previously been demonstrated to specify the left-right axis using the EGF-CFC co-receptors.
Highlights
Establishment of the left-right axis is important for positioning organs asymmetrically in the developing vertebrate-embryo
Our study suggests a putative molecular mechanism by which the initiation of the formation of Left right (L-R) axis might be established via the transcriptional regulation of Cryptic gene expression by Snail
For this study we retrieved the human Cryptic promoter sequence from Ensembl and identified putative transcription factors that might bind on this sequence, using the online database “ConSite”
Summary
Establishment of the left-right axis is important for positioning organs asymmetrically in the developing vertebrate-embryo. The EGF-CFC family member Cryptic, that functions as a co-receptor for some TGF-beta ligands, is developmentally expressed in higher mammals and mutations in the gene cause loss or change in left-right axis asymmetry. We understand the process of organ formation in some detail, we lack knowledge of the initial molecular events such as signaling and transcriptional regulation that are triggered towards organ positioning [2]. Even as the initial set of molecular events leading to asymmetric distribution of body organs is largely unknown, limited understanding of the L-R axis specification includes asymmetric expression of TGF-β family members Nodal and Lefty, along with the transcription factor Pitx on the prospective left side of the embryo [6,7,8].
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