(ii) To confirm that induced levels of MCPyV ST in i293-ST cells are representative of ST expression in the MCPyV-positive MCC cell lines, immunoblotting was performed using an MCPyV T-specific antibody comparing cell lysates from 1 105 cells of uninduced and induced i293-ST, MCC13, and MKL-1 cells
July 19, 2021
(ii) To confirm that induced levels of MCPyV ST in i293-ST cells are representative of ST expression in the MCPyV-positive MCC cell lines, immunoblotting was performed using an MCPyV T-specific antibody comparing cell lysates from 1 105 cells of uninduced and induced i293-ST, MCC13, and MKL-1 cells. of cellular proteins implicated in microtubule-associated cytoskeletal organization and dynamics. Intriguingly, we demonstrate that MCPyV ST expression promotes microtubule destabilization, leading to a motile and migratory phenotype. We further highlight the essential role of the microtubule-associated protein stathmin in MCPyV ST-mediated microtubule destabilization and cell motility and implicate the cellular phosphatase catalytic subunit protein phosphatase 4C (PP4C) in Ciproxifan maleate the regulation of this process. These findings suggest a possible molecular mechanism for the highly metastatic phenotype associated with MCC. IMPORTANCE Merkel cell polyomavirus (MCPyV) causes the majority of cases of Merkel cell carcinoma (MCC), an aggressive skin cancer with a high metastatic potential. However, the molecular mechanisms leading to virally induced cancer development have yet to be fully elucidated. In particular, no studies have investigated any potential link between the virus and the highly metastatic nature of MCC. We demonstrate that this MCPyV small tumor antigen (ST) promotes the destabilization of the host cell microtubule network, which leads to a more motile and migratory cell phenotype. We further show Ciproxifan maleate that MCPyV ST induces this process by regulating Ciproxifan maleate the phosphorylation status of the cellular microtubule-associated protein stathmin by its known association with the cellular phosphatase catalytic subunit PP4C. These findings highlight stathmin as a possible biomarker of MCC and as a target for novel antitumoral therapies. INTRODUCTION Merkel cell carcinoma (MCC) is an aggressive skin tumor (1). The reported cases of MCC have tripled in the past 20 years in both Europe and the United States (2), due to an increase in known risk factorsUV exposure, immune suppression, and increased age (1, 3). The cancer is usually characterized by significant incidence of local recurrence, early involvement of local lymph nodes, and distant metastasis (4). As such, MCC has a poor 5-year survival rate, due to its high propensity to metastasize (5). Merkel cell polyomavirus (MCPyV) is usually clonally integrated in 80% of MCC tumors (6). MCPyV encodes both large and small T antigens (LT and ST, respectively), which are regulatory proteins required for Ciproxifan maleate viral replication and tumorigenesis (6). MCPyV contamination and integration occur prior to expansion and metastasis of the tumor (7, 8), and truncation mutations of the LT gene are observed in the integrated genome rendering the virus replication defective (6). LT and ST are required for MCC cell survival and proliferation, as depletion of these T antigens leads to cell arrest and death of MCPyV-positive MCC cells (9). In contrast to simian virus 40 (SV40), MCPyV ST is sufficient to transform rodent cells to anchorage- and contact-independent growth and also induces serum-free proliferation of human cells (10). However, the exact contribution of ST to MCC cell growth is usually under debate as several ST depletion studies have shown differential dependence for MCC proliferation (11, 12). Recent analyses suggest that MCPyV ST is usually multifunctional in nature (13). MCPyV ST leads to the hyperphosphorylation of 4E-BP1, resulting in the deregulation of cap-dependent translation, (10), it targets the cellular ubiquitin ligase Ciproxifan maleate SCFFwb7, stabilizing MCPyV LT and several cellular oncoproteins (14), and also functions as an inhibitor of NF-B-mediated transcription (15). Although these interactions are attributed to either MCPyV ST-mediated cellular transformation or MCPyV replication processes, to date, no studies have investigated any potential link between MCPyV T antigen expression and the highly metastatic nature of MCC. This is of significant importance as dissemination and metastasis correlate with poor MCC survival Mmp28 rates (16). Despite the clinical importance, the molecular basis by which cancer cells acquire the capability to migrate from the primary tumor remains to be fully elucidated (17). What is clear is usually that cell motility, migration, and invasion are.