Co-factors of LIM domains (Clims) are transcriptional co-regulators that interact with LIM domain-containing transcription factors as well as certain other DNA-binding proteins. Clim2, a member of this gene family, is highly expressed in epithelial cells, including epithelial cells of the limbal and central cornea. To investigate the role of Clims, we generated transgenic mice expressing a dominant-negative Clim molecule (DN-Clim) under the control of the keratin 14 promoter (K14-DN-Clim mice). In corneal epithelium, the transgene expression causes decreased expression of adhesion molecule BP180 and defective hemidesmosomes, leading to detachment of corneal epithelium from the underlying stroma, which in turn causes blisters, wounds and an inflammatory response. After a period of epithelial thinning, the corneal epithelium undergoes differentiation to an epidermis-like structure. Together, these data indicate that Clim proteins play important roles in epithelial adhesion and the maintenance of cornea epithelial differentiation. The long term goal of our research is to understand how cornea epithelial cell differentiation is maintained. Our hypotheses in the current application are that a) Clims directly regulate the expression of hemidesmosome component BP180, and b) that through suppression of canonical Wnt signaling, Clims suppress epidermis-type differentiation in the corneal epithelium. We propose three Aims to investigate these hypotheses. In the first Aim, we will use in vivo confocal imaging and expression profiling to characterize the molecular changes occurring at the initiation and during progression of corneal abnormalities in the K14-DN-Clim mice. We expect to define gene expression signatures associated with specific pathological abnormalities, thus providing new ideas about pathogenesis and treatment of corneal disease. In the second Aim, we will use chromatin immunoprecipitation and transcription assays to define the molecular mechanisms whereby Clim regulate the expression of genes encoding components required for cornea epithelial adhesion. We expect to discover the DNA-binding protein with which Clim interact on the BP180 gene in cornea epithelial cells. In the third Aim, we will investigate the mechanisms whereby Clims maintain cornea differentiation and suppress epidermis-like differentiation of the corneal epithelium. We will define the role of wounding in this process and elicit the role of Clims in Wnt signaling, which is critical for corneal epithelial differentiation. This research is likely to give new insights into both impaired epithelial cell adhesion and altered corneal epithelial differentiation -- features which characterize several diseases of the cornea, some of which cause blindness.
There are several blistering diseases that affect the cornea, frequently leading to limbal stem cell deficiency. The K14-DN-Clim mice reproduce these features and may provide insights into the response of the cornea to such insults and lead to ideas about new drug targets. Squamous metaplasia and altered differentiation are also a hallmark of several corneal diseases that cause blindness;understanding the mechanisms whereby epidermis-like differentiation is prevented in the cornea is highly significant to public health.