Vaquero-Sedas María I, Vega-Palas Miguel A
Instituto de Bioquímica Vegetal y Fotosíntesis, CSIC-Universidad de Sevilla, IBVF (CSIC-US), E41092 Seville, Spain.
Int J Mol Sci. 2024 Jan 4;25(1):672. doi: 10.3390/ijms25010672.
Telomeres protect the ends of linear eukaryotic chromosomes from being recognized as DNA double-strand breaks. Two major protein complexes are involved in the protection of telomeres: shelterin and CST. The dysfunction of these complexes can challenge the function of telomeres and lead to telomere fusions, breakage-fusion-bridge cycles, and cell death. Therefore, monitoring telomere fusions helps to understand telomeres biology. Telomere fusions are often analyzed by Fluorescent In Situ Hybridization (FISH) or PCR. Usually, both methods involve hybridization with a telomeric probe, which allows the detection of fusions containing telomeric sequences, but not of those lacking them. With the aim of detecting both types of fusion events, we have developed a nested PCR method to analyze telomere fusions in . This method is simple, accurate, and does not require hybridization. We have used it to analyze telomere fusions in wild-type and mutant plants altered in CTC1, one of the three components of the Arabidopsis CST telomere capping complex. Our results show that null mutant plants display fusions between all telomeric regions present in Arabidopsis chromosomes 1, 3 and 5, thus highlighting the widespread end-capping protection achieved by CTC1.
端粒可保护线性真核染色体的末端不被识别为DNA双链断裂。有两种主要的蛋白质复合物参与端粒的保护：端粒保护蛋白复合体（shelterin）和CST。这些复合物的功能障碍会挑战端粒的功能，并导致端粒融合、断裂-融合-桥循环和细胞死亡。因此，监测端粒融合有助于理解端粒生物学。端粒融合通常通过荧光原位杂交（FISH）或聚合酶链反应（PCR）进行分析。通常，这两种方法都涉及与端粒探针杂交，从而能够检测包含端粒序列的融合，但无法检测缺乏端粒序列的融合。为了检测这两种类型的融合事件，我们开发了一种巢式PCR方法来分析[具体生物]中的端粒融合。该方法简单、准确，且无需杂交。我们已用它来分析野生型和在拟南芥CST端粒封端复合体的三个组分之一CTC1中发生改变的突变体植物中的端粒融合。我们的结果表明，缺失突变体植物在拟南芥1号、3号和5号染色体上存在的所有端粒区域之间都出现了融合，从而突出了CTC1所实现的广泛的末端封端保护作用。
