Alternative Splicing of Cyr61 Is Regulated by Hypoxia and Significantly Changed in Breast Cancer

Hypoxia is known to induce the transcriptional activation of pathways involved in angiogenesis, growth factor signaling, and tissue invasion and is therefore a potential key regulator of tumor growth. Cyr61 (cysteine rich 61) is a secreted matricellular protein with proangiogenic capabilities and is transcriptionally induced under hypoxic conditions. High expression levels of Cyr61 were already detected in various cancer types and linked to tumor progression and advanced stages in breast cancer. Besides hypoxia, there is some evidence that posttranscriptional pre-mRNA processing could be involved in the regulation of Cyr61 expression, but was thus far not investigated. We studied the expression pattern of Cyr61 mRNA and protein in breast cancer cell lines as well as in matched pairs of noncancerous breast tissue, preinvasive lesions, and invasive breast cancers, respectively. In addition, we analyzed the potential regulatory capability of hypoxia on Cyr61 expression by functional tissue culture experiments. Our study revealed a stage-dependent induction of Cyr61 mRNA and protein in breast cancer tumorigenesis and for the first time alternative splicing of the Cyr61 gene due to intron retention. Breast carcinogenesis was accompanied by a shift from an intron 3 retaining toward an intron 3 skipping mRNA phenotype consecutively leading to processing of the biological active Cyr61 protein. The functional analyses strongly emphasize that hypoxia serves as a specific inducer of alternative Cyr61 splicing toward the intron skipping mRNA isoform with potential biological consequences in tumor cells. [Cancer Res 2009;69(5):2082-90]