The major molecular mechanism responsible for specialized gene products in tissues appears to be differential gene expression, controlled primarily at the level of transcription (1) but there is growing evidence that posttranscriptional, translational and posttranslational control of selective gene expression is equally important for cellular diversity and maintenance of particular phenotypes. The long range goal of our research is to understand the molecular mechanism that controls differential gene expression during development and differentiation in human epidemias. Terminal differentiation of epidermal cells is associated with the sequential induction of a family differentiation-specific structural and regulatory proteins, or unique to keratinocytes, one of which is the crosslinking enzyme transglutaminase type I. We are defining cis-acting and trans-acting factors that regulate tissue-specific and differentiation-specific expression of the transglutaminase typeI (TGM1) gene as an important step in understanding the regulators that control epithelial differentiation. We propose to study regulatory mechanisms of differential gene expression using TGM1 gene as a model, because this is, to our knowledge, the only keratinocyte-specific gene whose expression is regulated by all three transcriptional, posttranscriptional and translational mechanisms of control. Identification and characterization of regulators operating at all three levels of genetic information flow from gene to functional protein would facilitate our understanding of mechanisms and biochemical pathways underlying development and differentiation. We will use molecular biology techniques, in vitro tissue culture monolayer and reconstituted skin models as well as in vivo study in transgenic mice to: 1. define cis- acting and trans-acting regulatory factors that control transcriptional activity of the TGM1 gene in keratinocyte- and differentiation-specific manner as well as factors responsible for modulatory response of keratinocytes to environmental variations. In particular study of promoters inhibition by oligonucleotide-directed triple-helix formation will be important in both basic science and clinical settings. 2. to determine whether TGM1 is regulated by activation of alternative promoter by alternative splicing of pre-mRNA at the posttranscriptional level and whether an observed novel transcript is related to altered epidermal physiology. 3. to determine the translatability of TGM1 transcript variations 4. to determine the role of the homeoprotein trans-acting factor that we isolated in the TGM1 gene expression. These studies will enable us to characterize some regulators of epidermal biology and identify steps at which differentiation and development of epidermis is controlled and may identify the levels at which the normal genetic information has gone awry in the diseased state of epidermis.
