Circular RNAs of UDP-Glycosyltransferase (UGT) Genes Expand the Complexity and Diversity of the UGT Transcriptome

Abstract

The human UDP-glycosyltransferase (UGT) gene superfamily generates 22 canonical transcripts coding for functional enzymes and also produces nearly 150 variant UGT transcripts through alternative splicing and intergenic splicing. In the present study, our analysis of circRNA databases identified backsplicing events that predicted 85 circRNAs from UGT genes, with 33, 11, and 19 circRNAs from UGT1A, UGT2B4, UGT8, respectively. Most of these UGT circRNAs were reported by one database and had low abundance in cell- or tissue-specific contexts. Using reverse-transcriptase polymerase chain reaction with divergent primers and cDNA samples from human tissues and cell lines, we found 13 circRNAs from four UGT genes: UGT1A (three), UGT2B7 (one), UGT2B10 (one), and UGT8 (eight). Notably, all eight UGT8 circRNAs contain open reading frames that include the canonical start AUG codon and encode variant proteins that all have the common 274-amino acidN-terminal region of wild-type UGT8 protein. We further showed that one UGT8 circRNA (circ_UGT8-1) was broadly expressed in human tissues and cell lines, resistant to RNase R digestion, and predominately present in the cytoplasm. We cloned five UGT8 circRNAs into the Zinc finger with KRAB and SCAN domains 1 vector and transfected them into HEK293T cells. All these vectors produced both circRNAsand linear transcripts with varying circular/linear ratios (0.17-1.14).Western blotting and mass spectrometry assays revealed that only linear transcripts and not circRNAs were translated. In conclusion, our findings of nearly 100 circRNAs greatly expand the complexity and diversity of the UGT transcriptome; however, UGT circRNAs are expressed at a very low level in specific cellular contexts, and their biologic functions remain to be determined. SIGNIFICANCE STATEMENT: The human UGT gene transcriptome comprises 22 canonical transcripts coding for functional enzymes and approximately 150 alternatively spliced and chimeric variant transcripts. The present study identified nearly 100 circRNAs from UGT genes, thus greatly expanding the complexity and diversity of the UGT transcriptome. UGT circRNAs were expressed broadly in human tissues and cell lines; however, most showed very low abundance in tissue- and cell-specific contexts, and therefore their biological functions remain to be investigated.