Torrado Mario, Iglesias Raquel, Nespereira Beatriz, Centeno Alberto, López Eduardo, Mikhailov Alexander T
Developmental Biology Group, Institute of Health Sciences, University of La Coruña, As Xubias s/n, 15006 La Coruña, Spain.
Gene. 2009 Jul 1;440(1-2):28-41. doi: 10.1016/j.gene.2009.03.017. Epub 2009 Mar 31.
The cardiac ankyrin repeat domain 1 protein (ANKRD1, also known as CARP) has been extensively characterized with regard to its proposed functions as a cardio-enriched transcriptional co-factor and stress-inducible myofibrillar protein. The present results show the occurrence of alternative splicing by intron retention events in the pig and human ankrd1 gene. In pig heart, ankrd1 is expressed as four alternatively spliced transcripts, three of which have non-excised introns: ankrd1-contained introns 6, 7 and 8 (i.e., ankrd1-i6,7,8), ankrd1-contained introns 7 and 8 (i.e., ankrd1-i7,8), and ankrd1 retained only intron 8 (i.e., ankrd1-i8). In the human heart, two orthologues of porcine intron-retaining ankrd1 variants (i.e., ankrd1-i8 and ankrd1-i7,8) are detected. We demonstrate that these newly-identified intron-retaining ankrd1 transcripts are functionally intact, efficiently translated into protein in vitro and exported to the cytoplasm in cardiomyocytes in vivo. In the piglet heart, both the intronless and intron-retaining ankrd1 mRNAs are co-expressed in a chamber-dependent manner being more abundant in the left as compared to the right myocardium. Our data further indicate co-upregulation of the ankrd1 spliced variants in myocardium in the porcine model of diastolic heart failure. Most significantly, we demonstrate that in vivo forced expression of recombinant intronless ankrd1 markedly increases the levels of intron-retaining ankrd1 variants (but not of the endogenous main transcript) in piglet myocardium, suggesting that ANKRD1 may positively regulate the expression of its own intron-containing RNAs in response to cardiac stress. Overall, our findings demonstrate that in cardiomyocytes ANKRD1 can exist in multiple isoforms which may contribute to the functional diversity of this factor in heart development and disease.
心肌锚蛋白重复结构域1蛋白（ANKRD1，也称为CARP），就其作为心脏富集转录辅因子和应激诱导肌原纤维蛋白的假定功能而言，已得到广泛研究。目前的结果显示，猪和人类ankrd1基因中存在内含子保留事件导致的可变剪接。在猪心脏中，ankrd1表达为四种可变剪接转录本，其中三种含有未切除的内含子：包含ankrd1内含子6、7和8（即ankrd1-i6,7,8），包含ankrd1内含子7和8（即ankrd1-i7,8），以及ankrd1仅保留内含子8（即ankrd1-i8）。在人类心脏中，检测到猪内含子保留ankrd1变体的两种直系同源物（即ankrd1-i8和ankrd1-i7,8）。我们证明，这些新鉴定的内含子保留ankrd1转录本功能完整，在体外能有效翻译成蛋白质，并在体内心肌细胞中输出到细胞质中。在仔猪心脏中，无内含子和内含子保留的ankrd1 mRNA均以腔室依赖性方式共表达，与右心肌相比，在左心肌中更为丰富。我们的数据进一步表明，在舒张性心力衰竭的猪模型中，ankrd1剪接变体在心肌中共同上调。最显著的是，我们证明，在体内强制表达重组无内含子ankrd1可显著增加仔猪心肌中内含子保留ankrd1变体的水平（但不增加内源性主要转录本的水平），这表明ANKRD1可能在心脏应激反应中正向调节其自身含内含子RNA的表达。总体而言，我们的研究结果表明，在心肌细胞中ANKRD1可以以多种异构体形式存在，这可能有助于该因子在心脏发育和疾病中的功能多样性。
