Abstract 1373 The inhibitory effect of miR-190a on GRHL-1 as a novel avenue for diagnosis and treatment of psoriasis

Psoriasis is an incurable auto-inflammatory chronic skin disease. Despite its common prevalence worldwide, it is extensively complex on the cellular, genomic, and genetic levels. Psoriasis is characterized by hyper-proliferation and dedifferentiation of keratinocytes. In a recently published work, we identified significant differences in the histone epigenetic modification marker H3K27Ac between psoriasis lesion skin and healthy or non-lesion skin, throughout the genome. A bioinformatics analysis of the epigenetic differential sites revealed enrichment for putative binding sites to the transcription factor GRHL. Many of these GRHL sites were found in regions of genes related to skin differentiation and development. Additionally, we observed a correlation between the epigenetic changes in these putative GRHL binding sites and alterations in the expression of nearby genes. We found that in psoriasis lesions, there is an elevation of expression of GRHL-1 and GRHL-3. MiR-190a-5p shows therapeutic potential for psoriasis by down-regulating GRHL-1. Preliminary data indicates a decrease in miR-190a expression in psoriasis lesions compared to healthy skin, reinforcing this possibility. MiR-190a expression has been implicated in a few cancers, acting as either a tumor-suppressor or oncogene, but there are no studies correlating miR-190a-5p with psoriasis. To investigate whether GRHL-1 is a biochemical target of miR-190a, we cloned the 3'UTR of GRHL-1 into a reporter plasmid, psiCHECK-II, and co-transfected with a plasmid expressing miR-190a. We used a reporter assay system, in which the 3'UTR of a specific mRNA is cloned downstream of a reporter renilla/luciferase gene. We found that increasing the amount of miR-190a decreases the expression of the reporter luciferase if attached to the 3'UTR of GRHL-1. We further confirmed this by generating a mutation at the putative miRNA binding site, which abolished this inhibitory effect on GRHL-1 expression. Our next steps include checking whether the over-expression of the miRNA decreases the endogenous targeted protein. This will be accomplished by transfecting cells with mimic miR-190a and analyzing the expression level of the endogenous protein through Western Blot analysis. Once we establish that miR-190a can indeed regulate the expression of GRHL-1, our subsequent studies will focus on examining the biological effects of miR-190a on keratinocytes. Specifically, we will explore its impact on keratinocyte proliferation and differentiation. Additionally, we will employ mRNA array analysis to examine the effect of over-expressing miR-190a on whole gene expression and assess how many of these affected genes are also influenced by GRHL-1 expression. These findings present significant opportunities for new avenues of diagnosis and treatment for psoriasis, and potentially other skin diseases, such as squamous cell carcinoma.