In order to identify membrane proteins whose expression is induced by X-ray irradiation, we developed an antibody (Ab)-directed strategy using a phage Ab library. X-Ray-irradiated cells were screened with a phage Ab library in the presence of a large excess of polyclonal Abs prepared against membrane proteins that are commonly present at the surface of both X-ray-irradiated and nonirradiated cells. After isolation of Ab that bound only to X-ray-irradiated cells, the antigen was identified using MS. Using this approach, we found that expression of LY6D is induced in MCF10A cells by X-ray irradiation. The induction of LY6D expression is triggered through a pathway regulated by ATM, CHK2 and p53. This method is a new Ab-directed proteomic strategy for analysis of membrane proteins, and is applicable to various biological phenomena in situations in which both target molecule-expressing cells and nonexpressing cells are available.
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