ИСТИНА

Recoverin is a Ca2+-binding protein normally localized in photoreceptor cells of the retina where it controls rhodopsin phosphorylation by rhodopsin kinase thereby participating in regulation of visual transduction. Recoverin was also recognized as paraneoplastic (onconeural) antigen since it can be aberrantly expressed in malignant tumors and in some cases this phenomenon is accompanied by generation of respective serum autoantibodies and development of paraneoplastic syndrome (cancer-associated retinopathy). A role of recoverin in cancer cells remained unspecified. Here, we attempted (1) to estimate prevalence of recoverin aberrant expression and generation of autoantibodies against the protein in various cancers, (2) to investigate the mechanisms, underlying the aberrant expression of recoverin in tumors and (3) to determine localization of recoverin in cancer cells in vitro. Immunohistochemical examination of the tumor samples revealed that recoverin is expressed in about 75% of lung cancers and in about 68% of patients with different subtypes of renal cell carcinoma. In non-small cell lung cancer, the frequency of recoverin positive tumor cells inversely correlated with the grade of differentiation while in renal tumors expression of the protein has tendency to correlate with tumor size. Using Western blotting, it was found that in renal cancers the frequency of recoverin autoantibodies is very low (<1% of cases), but they occur in about 20% patients with lung cancer. The presence of autoantibodies against recoverin in the serum of lung cancer patients is highly specific (98%), but only moderately sensitive (20%). Epigenetic and transcriptional analysis indicated that expression of recoverin in cancer cells (melanoma tissues and cell lines) is mediated by demethylation of the certain CpGs of the recoverin gene region overlapping the promoter up-stream of the first exon and the first exon itself. Consistently, the treatment of the cells with DNA methyltransferase inhibitor 5-aza-2-deoxycytidine reduced the calculated mean level of the recoverin gene methylation and increased the amount of the recoverin mRNA transcript. Light and electronic microscopy studies of retinoblastoma Y79 cells, taken as a model of recoverin expressing cancer cell line, revealed that recoverin is distributed as clusters located in the cytoplasm. Although membrane association is a distinctive feature of the retinal counterpart, no binding of recoverin to plasma or nuclear membrane of the cancer cells was observed. Instead, recoverin was found to associate with unidentified cytoplasmic structures likely different from endoplasmic reticulum since co-localization of the protein with ER marker calnexin was lower than 5%. Taken together, these data provide necessary basis for establishing of recoverin function in cancer cells and allow considering this protein as a potential diagnostic and/or prognostic marker for tumors if used in combination with other markers of cancer.