Increased activation of Ras in psoriatic lesions

Ras functions as an essential upstream regulator of growth-factor-receptor-coupled signal transduction pathways. Ras is converted from an inactive GDP-bound state to an active GTP-bound state in response to receptor activation. Thus, the ratio of GTP/GDP bound to Ras is a measure of its state of activation. Mutations that stabilize the GTP-bound form of Ras result in constitutive activation and cellular transformation. The most widely used method for measuring Ras activation utilizes [32P]PO4 to label cellular nucleotide pools and is therefore limited to use with cultured cells. We have modified and adapted an enzyme-based method for rapid, precise measurement of Ras-bound GTP and GDP in normal and psoriatic human skin. This method does not require radiolabeling of cellular nucleotides. In cultured fibroblasts, the enzymatic and [32P]PO4 incorporation methods yielded similar results. Application of the enzymatic method to human skin revealed that 6% of Ras was in the active GTP-bound state in normal skin, compared to 15.4% of Ras in psoriatic lesions. The total amount of Ras normalized to protein content was similar in normal and psoriatic skin. Enhanced activation of Ras is likely a critical mediator of the increased cell growth characteristic of psoriatic lesions.