Pathways for maintenance of telomeres and common fragile sites during DNA replication stress

Abstract

Oncogene activation during tumour development leads to changes in the DNA replication programme that enhance DNA replication stress. Certain regions of the human genome, such as common fragile sites and telomeres, are particularly sensitive to DNA replication stress due to their inherently 'difficult-to-replicate' nature. Indeed, it appears that these regions sometimes fail to complete DNA replication within the period of interphase when cells are exposed to DNA replication stress. Under these conditions, cells use a salvage pathway, termed 'mitotic DNA repair synthesis (MiDAS)', to complete DNA synthesis in the early stages of mitosis. If MiDAS fails, the ensuing mitotic errors threaten genome integrity and cell viability. Recent studies have provided an insight into how MiDAS helps cells to counteract DNA replication stress. However, our understanding of the molecular mechanisms and regulation of MiDAS remain poorly defined. Here, we provide an overview of how DNA replication stress triggers MiDAS, with an emphasis on how common fragile sites and telomeres are maintained. Furthermore, we discuss how a better understanding of MiDAS might reveal novel strategies to target cancer cells that maintain viability in the face of chronic oncogene-induced DNA replication stress.

Keywords: RAD52; alternative lengthening of telomeres; cancer; common fragile sites; homologous recombination.

Figure 1.

Key steps involved in the BIR pathway. BIR initiates from a double strand DNA end that has been resected to generate a 3′ single stranded DNA overhang (i). This overhang then invades into a homologous DNA duplex to form D-loop (ii) followed by DNA synthesis and D-loop migration and subsequent initiation of complementary strand synthesis (iii) [103].

Figure 2.

MiDAS at difficult-to-replicate loci. (a) Representative image of the detection of MiDAS in HeLa cells treated with low dose aphidicolin (APH). The ongoing DNA synthesis marked by EdU incorporation (red) can be seen in relation to the telomeric DNA ends (green). DNA is stained with DAPI (blue). (b) A current model for MiDAS [100]. The cartoon shows how a BIR-like process (figure 1) might occur at telomeres and CFSs when a stalled replication fork is broken and new DNA synthesis is activated (red). The fact that the process is RAD51-independent (which is unusual for BIR) suggests that perhaps the annealing of the broken arm of the fork occurs at a DNA structure that is already in an open conformation due to the presence of an R-loop or a DNA secondary structure.

Figure 3.

The mechanisms for counteracting replication stress that prevent genome instability. Replication stress at difficult-to-replicate loci such as CFSs and telomeres (indicated by stars on the chromosome arms) is initially dealt with via a canonical RAD51-dependent pathway before the cells enter into mitosis. When this pathway fails, a non-canonical, RAD51-independent, process (MiDAS) takes over to prevent genomic instability. APH, aphidicolin, HU, hydroxyurea.

Figure 4.

What happens when MiDAS fails? The consequences of MiDAS failure and progression through mitosis with unreplicated DNA could be not only the formation of mitotic aberrations such as anaphase bridges, lagging chromatin and chromosome breaks/gaps, but also genomic instability in the next G1 cell cycle of daughter cells. Mitotic anaphase bridges, which are classified as either chromatin bridges or ultra-fine bridges, are observed when cells attempt to segregate incompletely replicated or unresolved DNA structures. This can lead to the daughter cells acquiring an incorrect chromosome number/structure, to the formation of micronuclei, and to the formation of so-called 53BP1 nuclear bodies in the daughter G1 cells.