Engineered Proteins for Controlling Gene Expression
This chapter describes the potential uses of natural transcriptional regulators, enhancement of these regulators through molecular engineering, and the engineering of entirely synthetic transcription factors for targeted gene regulation. HIF-1 is a heterodimeric transcription factor composed of two basic helix-loop-helix (bHLH) proteins, HIF-1α and HIF-1β. HIF-1β is constitutively expressed in an active form in the nucleus of oxygen-sensing cells and HIF-1α is highly inducible by hypoxia, primarily through post-translational regulation. Under normoxic conditions, HIF-1α is rapidly degraded through hydroxylation of proline residues in the N- and C-terminal oxygen-dependent degradation domains (NODDD and CODDD). This post-translational modification is mediated by a family of three HIF-1α prolyl hydroxylases that use oxygen as a substrate such that enzymatic activity is tightly regulated by oxygen concentration. Hydroxylated proline residues are recognized by the von Hippel Lindau tumor suppressor (VHL), which targets HIF-1α for proteosomal proteolysis via ubiquitin ligation. Artificial transcription factors based on zinc finger proteins are engineered to regulate a variety of genes relevant to regenerative medicine such as a zinc finger transcription factor is designed to regulate the gene for vascular endothelial growth factor (VEGF). VEGF is known to stimulate the formation of new blood vessels necessary for wound healing and the repair of injured cardiovascular tissues.
