Department of Biomedical Sciences, School of Medicine and Health Sciences, University of North Dakota, 501 North Columbia Road, Grand Forks, ND 58202, USA.
G3 (Bethesda). 2024 Sep 4;14(9). doi: 10.1093/g3journal/jkae165.
Poly(ADP-ribose) polymerase 1 (PARP1) is a highly conserved nuclear protein in multicellular organisms that by modulating chromatin opening facilitates gene expression during development. All reported Parp1 null knockout mouse strains are viable with no developmental anomalies. It was believed that functional redundancy with other PARP family members, mainly PARP2, explains such a controversy. However, while PARP2 has similar catalytic domain to PARP1, it lacks other domains, making the absence of developmental problems in Parp1 mice knockouts unlikely. Contrary to prior assumptions, in our analysis of the best-investigated Parp1 knockout mouse strain, we identified persistent mRNA expression, albeit at reduced levels. Transcript analysis revealed an alternatively spliced Parp1 variant lacking exon 2. Subsequent protein analysis confirmed the existence of a truncated PARP1 protein in knockout mice. The decreased level of poly(ADP-ribose) (pADPr) was detected in Parp1 knockout embryonic stem (ES) cells with western blotting analysis, but immunofluorescence staining did not detect any difference in distribution or level of pADPr in nuclei of knockout ES cells. pADPr level in double Parp1 Parg mutant ES cells greatly exceeded its amount in normal and even in hypomorph Parg mutant ES cells, suggesting the presence of functionally active PARP1. Therefore, our findings challenge the conventional understanding of PARP1 depletion effects.
多聚（ADP-核糖）聚合酶 1（PARP1）是一种在多细胞生物中高度保守的核蛋白，通过调节染色质的开放，促进发育过程中的基因表达。所有报道的 Parp1 基因敲除小鼠品系都是存活的，没有发育异常。人们认为，与其他 PARP 家族成员（主要是 PARP2）的功能冗余解释了这种争议。然而，虽然 PARP2 具有与 PARP1 相似的催化结构域，但它缺乏其他结构域，这使得 Parp1 基因敲除小鼠中不存在发育问题的可能性不大。与之前的假设相反，在我们对研究最充分的 Parp1 基因敲除小鼠品系的分析中，我们发现了持续的 mRNA 表达，尽管水平降低。转录分析显示存在一种缺乏外显子 2 的 PARP1 剪接变体。随后的蛋白质分析证实了在敲除小鼠中存在截短的 PARP1 蛋白。Western blot 分析检测到 Parp1 基因敲除胚胎干细胞（ES 细胞）中聚（ADP-核糖）（pADPr）水平降低，但免疫荧光染色未检测到敲除 ES 细胞中 pADPr 在核内的分布或水平有任何差异。双 Parp1 Parg 突变型 ES 细胞中的 pADPr 水平大大超过了正常 ES 细胞甚至低表达型 Parg 突变型 ES 细胞中的水平，表明存在功能活跃的 PARP1。因此，我们的研究结果对 PARP1 耗竭效应的传统认识提出了挑战。
