Abstract:
The CXCL12 gene produces a series of transcript variants through alternative splicing at the 3\' end of its pre-mRNA. This study explores the biological activities of these alternative transcripts and the mechanisms involved in the regulation of CXCL12 transcription and RNA splicing. We identified a \"GA\" insertion mutation in the region of CXCL12α DNA encoding the conserved 3\'UTR. This variant transcript was named CXCL12-3\'GA+. The mutation occurred at a frequency of 13.2% in healthy Chinese individuals. However, its frequency in healthy Caucasians was 22.6%, significantly higher than what was observed in the Chinese. Genomic analysis indicated that the GA+ mutation likely encodes a G-quadruplex structure in close proximity to a cluster of important AU-rich elements (AREs) that are well-established regulators of mRNA stability at the 3\'UTR. Experiments using molecular constructs encoding the 3\'UTR of CXCL12 revealed that the GA+ allele can significantly increase gene expression compared to the WT allele. Further studies uncovered that the WT allele was associated with the production of a 225-bp minor transcript isoform (MTI) through alternative splicing resulting in the deletion of exon 2. ARMS-PCR using samples collected from cultured PBMCs of WT/GA+ genotype carriers indicated that the GA+ allele was preferentially transcribed compared to the WT allele. In summary, the study demonstrates that a GA insertion in the region encoding the 3\'UTR of CXCL12α may affect gene expression through alternative mRNA splicing. This finding provides a basis for understanding how multiple elements in the sequence encoding the 3\'UTR of the CXCL12 gene regulates its transcription and may lead to insights about diseases involving abnormal CXCL12α expression.
摘要:
CXCL12基因通过在其前mRNA的3'末端的选择性剪接产生一系列转录变体。本研究探讨了这些替代转录本的生物学活性以及参与CXCL12转录和RNA剪接调控的机制。我们在编码保守的3'UTR的CXCL12αDNA区域中鉴定了一个“GA”插入突变。该变体转录物被命名为CXCL12-3\'GA+。该突变在健康的中国个体中以13.2%的频率发生。然而,其在健康白种人中的频率为22.6%,明显高于在中国观察到的。基因组分析表明,GA突变可能编码G-四链体结构,紧邻重要的富含AU的元件(ARE)簇,这些元件是3'UTR上mRNA稳定性的公认调节剂。使用编码CXCL12的3'UTR的分子构建体的实验表明,与WT等位基因相比,GA+等位基因可以显著增加基因表达。进一步的研究发现,WT等位基因与通过可变剪接导致外显子2缺失的225bp次要转录同种型(MTI)的产生有关。使用从WT/GA+基因型携带者的培养PBMC收集的样品的ARMS-PCR表明,与WT等位基因相比,GA+等位基因优先转录。总之,该研究表明,在CXCL12α的3'UTR编码区域中插入GA可能通过选择性mRNA剪接影响基因表达。这一发现为理解编码CXCL12基因3'UTR的序列中的多个元件如何调节其转录提供了基础,并可能导致有关涉及异常CXCL12α表达的疾病的见解。