Proteomic analysis of high-molecular-weight protein polymers in a doxorubicin-resistant breast-cancer cell line

We recently reported that increased transglutaminase 2 (TGase 2) expression correlates with increased resistance to the cancer drug doxorubicin in breast-cancer cell lines. Interestingly, high-molecular-weight (HMW) proteins also increased with increased TGase 2 expression in the drug-resistant cell lines. TGase 2 is likely to be responsible for the formation of HMW proteins, because TGase 2 catalyzes cross-linking between proteins. Although the role of the HMW proteins is unclear, we demonstrated that TGase 2 inhibition increases drug sensitivity in breast-cancer cells. Herein we find that TGase 2 inhibition by cystamine dramatically reduces the level of HMW proteins. Identification of the HMW proteins may suggest the mechanism of cancer drug resistance associated with aberrant TGase 2 function. To explore the identities of HMW proteins, we performed in-gel tryptic digestions of unresolved HMW proteins and analyzed the resulting peptides using LC-MALDI-MS/MS. Most of the identified proteins were associated with gene regulation, such as polyadenylate-binding proteins, translation initiation factors, and ribonucleoproteins. This finding suggests that TGase 2 may participate in gene regulation, in addition to its role in cell adhesion.