Analysis of transcripts from alternative PRKAR1B gene promoters in colorectal cancer

Abstract

Background/Objectives: The transcriptional regulation of PRKAR1B is controlled by alternative promoters, and previous in silico analysis has indicated their differential activity in colon and rectal cancer tissue in comparison to normal gut mucosa. The aim of this study was to investigate PRKAR1B promoters and transcripts potentially involved in cancer. Methods: The sequences of PRKAR1B alternative promoters were retrieved from Ensembl database: promoter A 752209 and promoter B 767287 bases upstream from the translation start site. Bioinformatic tools Alggen, AliBaba, CiiiDER, and TFBIND were used to predict binding of transcriptional regulators. Primer extension assay was performed on RNA isolated from malignant colon cell lines using an oligonucleotide probe binding to the sequence at the exon2/exon3 junction common for all PRKAR1B transcripts. Results: Based on analyzed elements, both PRKAR1B promoters were found to have atypical structure. According to the prediction, promoter A that encodes transcript PRKAR1B-201 binds several factors involved in cell proliferation, while promoter B that encodes transcript PRKAR1B-203 binds mostly pro-apoptotic factors. In primer extension experiments, a single signal corresponding to the transcript PRKAR1B-212 was observed in malignant cells. Conclusion: The differential activity of alternative PRKAR1B promoters in colorectal cancer can be explained by in silico results, predicting that promoter sequences bind sets of transcriptional regulators with opposing roles. However, experiments point to the transcript unrelated to either of the investigated promoters as potential cancer biomarker and it should be further characterized.