Targeted Proteomic Analysis of Small GTPases in Radioresistant Breast Cancer Cells

Radiation therapy benefits more than 50% of all cancer
patients
and cures 40% of them, where ionizing radiation (IR) deposits energy
to cells and tissues, thereby eliciting DNA damage and resulting in
cell death. Small GTPases are a superfamily of proteins that play
critical roles in cell signaling. Several small GTPases, including
RAC1, RHOB, and RALA, were previously shown to modulate radioresistance
in cancer cells. However, there is no systematic proteomic study on
small GTPases that regulate radioresistance in cancer cells. Herein,
we applied a high-throughput scheduled multiple-reaction monitoring
(MRM) method, along with the use of synthetic stable isotope-labeled
(SIL) peptides, to identify differentially expressed small GTPase
proteins in two pairs of breast cancer cell lines, MDA-MB-231 and
MCF7, and their corresponding radioresistant cell lines. We identified
7 commonly altered small GTPase proteins with over 1.5-fold changes
in the two pairs of cell lines. We also discovered ARFRP1 as a novel
regulator of radioresistance, where its downregulation promotes radioresistance
in breast cancer cells. Together, this represents the first comprehensive
investigation about the differential expression of the small GTPase
proteome associated with the development of radioresistance in breast
cancer cells. Our work also uncovered ARFRP1 as a new target for enhancing
radiation sensitivity in breast cancer.