Departments of Medicine, David Geffen School of Medicine, University of California, Los Angeles, CA, USA.
Departments of Human Genetics, David Geffen School of Medicine, University of California, Los Angeles, CA, USA.
Nucleus. 2023 Dec;14(1):2262308. doi: 10.1080/19491034.2023.2262308. Epub 2023 Sep 27.
The knockout mouse () created by Sullivan and coworkers in 1999 has been widely used to examine lamin A/C function. The knockout allele contains a deletion of intron 7-exon 11 sequences and was reported to be a null allele. Later, Jahn and coworkers discovered that the mutant allele produces a 54-kDa truncated lamin A and identified, by RT-PCR, a cDNA containing exon 1-7 + exon 12 sequences. Because exon 12 encodes prelamin A's motif, the mutant lamin A is assumed to be farnesylated. In the current study, we found that the truncated lamin A in mouse embryonic fibroblasts (MEFs) was predominantly nucleoplasmic rather than at the nuclear rim, leading us to hypothesize that it was not farnesylated. Our study revealed that the most abundant transcripts in MEFs contain exon 1-7 but not exon 12 sequences. Exon 1-7 + exon 12 transcripts were detectable by PCR but in trace amounts. We suspect that these findings explain the nucleoplasmic distribution of the truncated lamin A in MEFs, and subsequent cell transduction experiments support this suspicion. A truncated lamin A containing exon 1-7 sequence was nucleoplasmic, whereas a lamin A containing exon 1-7 + exon 12 sequences was located along the nuclear rim. Our study explains the nucleoplasmic targeting of truncated lamin A in MEFs and adds to our understanding of a commonly used strain of mice.
1999 年，由沙利文（Sullivan）及其同事创建的基因敲除鼠（）已被广泛用于研究核纤层蛋白 A/C 的功能。该敲除等位基因缺失了内含子 7-外显子 11 序列，据报道为无功能等位基因。后来，扬（Jahn）及其同事发现该突变等位基因产生了一个 54kDa 的截断核纤层蛋白 A，并通过 RT-PCR 鉴定了一个包含外显子 1-7+外显子 12 序列的 cDNA。由于外显子 12 编码前核纤层蛋白 A 的棕榈酰化基序，因此推测突变的核纤层蛋白 A 被棕榈酰化。在本研究中，我们发现基因敲除鼠的胚胎成纤维细胞（MEFs）中的截断核纤层蛋白 A 主要位于核质中，而不是核周缘，这使我们假设其未被棕榈酰化。我们的研究表明，在基因敲除鼠的 MEFs 中，最丰富的 转录本包含外显子 1-7，但不包含外显子 12 序列。通过 PCR 可以检测到外显子 1-7+外显子 12 转录本，但含量很少。我们怀疑这些发现解释了截断核纤层蛋白 A 在基因敲除鼠 MEFs 中的核质分布，随后的细胞转导实验支持了这一假设。含有外显子 1-7 序列的截断核纤层蛋白 A 位于核质中，而含有外显子 1-7+外显子 12 序列的核纤层蛋白 A 则位于核周缘。我们的研究解释了截断核纤层蛋白 A 在基因敲除鼠 MEFs 中的核质靶向，并增加了我们对常用基因敲除鼠品系的理解。
