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Extrachromosomal telomeric circles are commonly invoked as important players in telomere maintenance, but their origin has remained elusive. Using electron microscopy analysis on purified telomeres we show that, apart from known structures, telomeric repeats accumulate internal loops (i-loops) that occur in the proximity of nicks and single-stranded DNA gaps. I-loops are induced by single-stranded damage at normal telomeres and represent the majority of telomeric structures detected in ALT (Alternative Lengthening of Telomeres) tumor cells. Our data indicate that i-loops form as a consequence of the exposure of single-stranded DNA at telomeric repeats. Finally, we show that these damage-induced i-loops can be excised to generate extrachromosomal telomeric circles resulting in loss of telomeric repeats. Our results identify damage-induced i-loops as a new intermediate in telomere metabolism and reveal a simple mechanism that links telomere damage to the accumulation of extrachromosomal telomeric circles and to telomere erosion.

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Chromosome instability (CIN) is the most common form of genome instability and is a hallmark of cancer. CIN invariably leads to aneuploidy, a state of karyotype imbalance. Here, we show that aneuploidy can also trigger CIN. We found that aneuploid cells experience DNA replication stress in their first S-phase and precipitate in a state of continuous CIN. This generates a repertoire of genetically diverse cells with structural chromosomal abnormalities that can either continue proliferating or stop dividing. Cycling aneuploid cells display lower karyotype complexity compared to the arrested ones and increased expression of DNA repair signatures. Interestingly, the same signatures are upregulated in highly-proliferative cancer cells, which might enable them to proliferate despite the disadvantage conferred by aneuploidy-induced CIN. Altogether, our study reveals the short-term origins of CIN following aneuploidy and indicates the aneuploid state of cancer cells as a point mutation-independent source of genome instability, providing an explanation for aneuploidy occurrence in tumors.

Plant cells are endowed with two distinct DNA polymerases [ 1,2] whose properties closely resemble those of the DNA polymerases cr and y present in animal cells [3,4]. The plant DNA polymerases have consequently been named o-like [ 1 ] and y-like [2].The B-like DNA polymerase activity is the most abundant in cultured plant cells [l ] and responds to changes in the rate of cell multiplication, whereas experiments with spinach leaves have shown that the y-like DNA polymerase is present in the chloroplast 121.A DNA polymerase has also been isolated from the mitochondria of wheat embryos [S]. Spinach mitochondria may also contain a DNA polymerase whose properties are partially different from those of the y-like DNA polymerase isolated from the chloroplasts of the same cells and are similar to those of the wheat embryo enzyme (unpublished).However, no evidence is available as yet on the existence in plant cells of a DNA repair enzyme similar to the DNA polymerase p of mammalian cellsBy analogy with animal cells, the assignment of functions to the DNA polymerases in plant cells is hampered by the lack of conditional mutants defective in DNA synthesis. Thus, we approach this problem by exploiting the properties of aphidicolin and of ethidium bromide.Aphidicolin We now describe the effect of aphidicolin and of ethidium bromide on the synthesis of DNA in the nucleus, c~oroplast and mitochondrion of cultured rice cells, as assessed by autoradiography at light and electron microscopy. The results show that aphidicolin specifically prevents the synthesis of nuclear DNA, while it has no effect on the synthesis of the organellar DNA, the latter being specifically affected only by ethidium bromide. This, together with the previous demonstration that only the a-like DNA polymerase is inhibited in vitro by aphidicolin [8], proves that the plant a-like DNA polymerase plays an essential role in the replication of nuclear DNA, and that this enzyme is not involved in the replication of plastid and mitochondriai DNA. Thus, the replication of the plant organellar DNA requires different DNA polymerases, the aphidicolin-resistant DNA polymerases present in the chloroplast [2] and in the plant mitochondrion [S J being the best candidates for this function.