Alternative Lengthening of Telomeres (ALT) Mechanism
Telomere instability results in senescence and carcinogenesis. Telomere length in cancer cells can be maintained by telomerase or alternative lengthening of telomeres (ALT). It is promising to target telomere maintenance mechanisms for cancer therapy. ALT mechanism is particularly prevalent in tumors of mesenchymal origin, including sarcomas, brain tumors, and pancreatic neuroendocrine tumors, etc. ALT-type cancers are difficult to treat and are associated with poor prognosis. The knowledge gap is to understand how ALT is initiated and maintained. In recent years, we developed quantitative approaches to study the molecular mechanisms. We reported that replication-associated damage triggers ALT (Zhang*, PLoS Genetics, 2019). Intriguingly, BLM helicase generates long 5’-Flaps at unligated Okazaki fragments during lagging strand replication to elicit a robust damage response that orchestrates ALT (Jiang*, Zhang*, Mol Cell, 2024, *equal contribution). We developed the first methodologies to purify telomere DNA damage response proteome and demonstrated that ALT-associated break-induced replication combines long-tract homology-directed repair synthesis with template switching to bypass replication obstacles, contributing to ALT telomere lengthening and rearrangement (Zhang, Nature, 2023). These findings revealed the key events and molecules that initiate and maintain ALT.
ALT telomeres harbor myriad forms of DNA damage responses for noncanonical recombination-dependent DNA synthesis. My previous studies implicate extensive PCNA-ubiquitination and ATRX deficiency in directing the assembly of DNA damage tolerance and recombination mechanisms during ALT. The big question is to understand how replisome and nucleosome alterations orchestrate communication between DNA replication and repair machinery to maintain ALT.
Centromere integrity maintenance
Centromeres are chromosomal regions required for proper chromosome segregation in mitosis and meiosis. Centromere integrity ensures accurate genetic information inheritance and prevents aneuploidy. Centromere instability leads to genome rearrangement, mutations, and chromosome instability, which contributes to the initiation and evolution of many tumor types, and genetic diseases, like ICF syndrome (Immunodeficiency, Centromeric region instability, Facial anomalies syndrome). However, centromeres are characterized as difficult-to-replicate regions and DNA damage hotspots. Homology-directed repair occurs frequently at the centromere regardless of DNA damage. The histone H3 variant CENP-A defines the unique epigenetic feature of centromeres, playing a central role in marking centromeres and establishing kinetochores. The exclusive centromere features co-direct physiological roles in replication, damage repair, and chromosome separation. We hypothesize that replication-coupled repair at the centromere assembles multiple DNA repair mechanisms for efficient resolution of lesions that constrain canonical homology-directed repair with sister chromatids. We aim to define the spatiotemporal regulation of centromeric homology-directed repair in different cell cycles and explore the unique epigenetic regulation of replication-coupled repair at centromeres.
Genome integrity maintenance within repetitive sequences
Genome instability in repetitive sequences results in numerous diseases, including cancers, neurodegeneration diseases, ageing and other syndromes. Complicated secondary structures and heterochromatin-like chromatin at the repetitive DNA sequences challenge the DNA replication machinery. These genome regions are characterized as difficult-to-replicate regions and DNA damage hotspots, which are prone to homology-directed repair. Break-induced replication occurs not only at ALT telomeres but also widely in the genome, especially repetitive sequences, like common fragile sites, centromere, short tandem repeats, microsatellites, etc. Consequently, break-induced replication results in genome rearrangement, translocation, and copy number gain/loss. We are dedicated to establishing new approaches to study genome integrity maintenance within repetitive sequences. We aim to address the following questions: How are repetitive sequences replicated? How are repetitive sequences repaired? How are error-free homologous recombination and error-prone break-induced replication being regulated?