New challenges in pseudopodial proteomics by a laser-assisted cell etching technique

Pseudopodia are ventral membrane protrusions that extend toward higher concentrations of chemoattractants and play key roles in cell migration and cancer cell invasion. Cancers, including carcinoma and sarcoma, become life threatening when they invade surrounding structures and other organs. Understanding the molecular basis of invasiveness is important for the elimination of cancers. Thus, determining the pseudopodial composition will offer insights into the mechanisms underlying tumor cell invasiveness and provide potential biomarkers and therapeutic targets. Pseudopodial composition has been extensively investigated by using proteomic approaches. A variety of modalities, including gel-based and mass spectrometry-based methods, have been employed for pseudopodial proteomics. Our research group recently established a novel method using excimer laser pulses to selectively harvest pseudopodia, and we successfully identified a number of new pseudopodial constituents. Here, we summarized the conventional proteomic procedures and describe our new excimer laser-assisted method, with a special emphasis on the differences in the methods used to isolate pseudopodia. In addition, we discussed the theoretical background for the use of excimer laser-mediated cell ablation in proteomic applications. Using the excimer laser-assisted method, we showed that alpha-parvin, an actin-binding adaptor protein, is localized to pseudopodia, and is involved in breast cancer invasiveness. Our results clearly indicate that excimer laser-assisted cell etching is a useful technique for pseudopodial proteomics. This article is part of a Special Issue entitled: Medical Proteomics. (C) 2014 Elsevier B.V. All rights reserved.