IGF-II

Description

Insulin-like growth factor II (IGF-II) functions critically in development and disease. The study of the murine IGF-II gene, Igf2 , has provided significant insight into epigenetic mechanisms of regulation. The parentally-imprinted expression of Igf2 and the oppositely imprinted H19 gene are determined by the reciprocal binding of CTCF, a chromatin insulator protein. This regulatory mechanism served as a model for allele-specific repression of the X chromosome in female mammals. X chromosome inactivation (XCI) is regulated by two non-coding, overlapping RNAs: Xist , which is expressed from the X chromosome destined for inactivation, and Tsix , which is expressed from the X that will remain active. A GC-rich region downstream of the Tsix promoter, which was demonstrated previously to control both random and imprinted XCI, was found to contain putative CTCF binding sites. Because CTCF functions as a chromatin insulator that can prevent enhancers from acting on promoters, I cloned regions of Tsix that contained putative binding sites and tested them in an in vitro enhancer-blocking assay. The putative CTCF sites blocked the activation of a drug-resistance reporter gene from a locus control region (LCR), which acts as a transcriptional enhancer, and demonstrated that the 5' Tsix region functions as a chromatin insulator. IGF-II is upregulated by sonic hedgehog (Shh), a developmental morphogen that is misexpressed in various disorders. Shh was identified recently as an angiogenic factor in part through its ability to upregulate vascular endothelial growth factor (VEGF); however, the mechanisms involved in this interaction had not been explored. I determined that the primary mediator of Shh signaling, the transcription factor Gli, did not activate VEGF expression. This suggested that Shh did not directly upregulate VEGF transcriptionally. I determined that many effects of Shh, such as cell differentiation and proliferation, were augmented by IGF-II. The synergism between Shh and IGF-II was attributed to enhanced Gli activation by IGF-II. Furthermore, IGF-II upregulated expression of an in vitro VEGF reporter, and induced VEGF protein production in murine epidermal cells. This induction was dependent on PI3-kinase, and may involve the cooperative activity of NFκB and Sp1.

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