Institute of Animal Husbandry, Heilongjiang Academy of Agricultural Sciences, Harbin 150086, China.
College of Animal Science and Technology, Northeast Agricultural University, Harbin 150030, China.
Genes (Basel). 2022 Jul 22;13(8):1304. doi: 10.3390/genes13081304.
The cAMP response element-binding protein (CREB), a basic leucine zipper transcription factor, is involved in the activation of numerous genes in a variety of cell types. The gene is rich in alternative splicing (AS) events. However, studies on the AS of genes in pigs are limited, and few reports have compared the roles of isoforms in activating gene expression. Here, five AS transcripts, V1-5, were characterized by RT-PCR and two, V3 and V5, were new identifications. Both V1 and V2 have all the functional domains of the CREB protein, with similar tissue expression profiles and mRNA stability, suggesting that they have similar roles. The transcriptional transactivation activities of four isoforms encoding complete polypeptides were analyzed on the expression of the B-cell CLL/lymphoma 2-like protein 2 and the poly (A)-binding protein, nuclear 1 genes with a dual-luciferase reporter system, and differential activities were observed. Both V1 and V2 have promoting effects, but their roles are gene-specific. V3 has no effect on the promoter of the two genes, while V4 functions as a repressor. The mechanisms underlying the differential roles of V1 and V2 were analyzed with RNA-seq, and the genes specifically regulated by V1 and V2 were identified. These results will contribute to further revealing the role of CREB and to analyzing the significance of AS in genes.
环磷酸腺苷反应元件结合蛋白（CREB）是一种碱性亮氨酸拉链转录因子，参与多种细胞类型中许多基因的激活。该基因富含可变剪接（AS）事件。然而，关于猪中基因的 AS 研究有限，很少有报道比较同工型在激活基因表达中的作用。在这里，通过 RT-PCR 对 5 种 AS 转录本（V1-5）进行了表征，其中 2 种（V3 和 V5）是新发现的。V1 和 V2 都具有 CREB 蛋白的所有功能结构域，组织表达谱和 mRNA 稳定性相似，表明它们具有相似的作用。通过双荧光素酶报告基因系统分析了编码完整多肽的四个同工型对 B 细胞 CLL/淋巴瘤 2 样蛋白 2 和多聚（A）结合蛋白核 1 基因表达的转录激活活性，并观察到了差异活性。V1 和 V2 都有促进作用，但它们的作用是基因特异性的。V3 对两个基因的启动子没有影响，而 V4 则作为抑制剂。通过 RNA-seq 分析了 V1 和 V2 差异作用的机制，并鉴定了 V1 和 V2 特异性调控的基因。这些结果将有助于进一步揭示 CREB 的作用，并分析 AS 在基因中的意义。
