70-kDa Heat Shock Proteins (HSPA/HSP70) are chaperones playing a central role in the proteostasis control mechanisms. Their basal expression can be highly elevated as an adaptive response to environmental and pathophysiological stress conditions. HSPA2, one of poorly characterised chaperones of the HSPA/HSP70 family, has recently emerged as epithelial cells differentiation-related factor. It is also commonly expressed in cancer cells, where its functional significance remains unclear. Previously, researchers at the Maria Sklodowska-Curie National Research Institute of Oncology have found that proteotoxic stress provokes a decrease in HSPA2 levels in cancer cells. In the present study the researchers found that proteasome inhibition-related loss of HSPA2 from cancer cells neither is related to a block in the gene transcription nor does it relate to increased autophagy-mediated disposals of the protein. Proteotoxic stress stimulated extracellular release of HSPA2 in extracellular vesicles (EVs). Interestingly, EVs containing HSPA2 are also released by non-stressed cancer and normal cells. In human urinary EVs levels of HSPA2 were correlated with the levels of TSG101, one of the main EVs markers. The researchers conclude that HSPA2 may constitute basic components of EVs. Nevertheless, its specific role in EVs and cell-to-cell communication requires further investigation.
Immunodetection of HSPA2, EVs markers, and prostate-related marker (PSMA)
in urinary EVs isolated using antibody-conjugated beads
(a) Beads were coated with the mixture of anti-CD63, anti-CD81 and anti-CD9 antibodies in equal proportions or (b) with anti-PSMA antibody. Urine samples from one male healthy donor (HD) and two prostate cancer patients (PC1; PC2) were tested. A sample with PBS and coated beads was used as a control for non-specific antibody interaction during Western blot (WB) procedure. Antibody fragments detached from the coated beads during samples preparation for WB were visualized with Ponceau S staining as a control of the sample loading and electrotransfer performance. Membranes were cut into fragments according to the proteins’ molecular weight. For chemiluminescent signal detection X-ray film was used.