CTC1-STN1-TEN1 controls DNA break repair pathway choice via DNA end resection blockade.

Antagonistic activities of the 53BP1 axis and the tumor suppressor BRCA1-BARD1 determine whether DNA double-strand breaks (DSBs) are repaired by end joining or homologous recombination. We show that the CTC1-STN1-TEN1 (CST) complex, a central 53BP1 axis component, suppresses DNA end resection by EXO1 and the BLM-DNA2 helicase-nuclease complex but acts by distinct mechanisms in restricting these entities. Whereas BRCA1-BARD1 alleviates the CST-imposed EXO1 blockade, it has little effect on BLM-DNA2 restriction.

Structural basis for Rad54- and Hed1-mediated regulation of Rad51 during the transition from mitotic to meiotic recombination.

Rad51 catalyzes the DNA pairing reactions that take place during homologous recombination (HR), and HR must be tightly regulated to ensure physiologically appropriate outcomes. Rad54 is an ATP-dependent DNA motor protein that stimulates Rad51 activity during mitosis. In meiosis Rad51 is downregulated by the protein Hed1, which blocks Rad54 binding to Rad51, and allows Dmc1 to function as the active recombinase.

Biochemical Mechanisms of Genetic Recombination and DNA Repair.

Genetic recombination involves the exchange of genetic material between homologous sequences of DNA. It is employed during meiosis in sexually reproducing organisms or in somatic cells to accurately repair toxic DNA lesions like double-strand breaks and stalled replication forks. In these separate roles, recombination drives genetic diversity by enabling reshuffling of parental genetic information while also serving as a molecular safeguard against the deleterious effects of gross chromosomal rearrangements or mutagenic insults arising for either endogenous or exogenous reasons.

Single-Molecule Visualization of BLM-DNA2-Mediated DNA End Resection Using DNA Curtains.

Homologous recombination (HR) is the principal pathway undertaken by a cell for the error-free repair of DNA double-strand breaks that are frequently encountered by the cell. HR can be initiated at the sites of DNA double-strand breaks by generating long stretches of single-stranded 3' DNA overhang through a process called DNA end resection. In one DNA end resection pathway, this is achieved via the concerted effort of specialized machinery involving the RecQ family helicase BLM, the helicase/endonuclease DNA2, and a single-strand DNA binding protein complex RPA.

Lineage-specific amino acids define functional attributes of the protomer-protomer interfaces for the Rad51 and Dmc1 recombinases.

Most eukaryotes possess two Rad51/RecA family DNA recombinases that are thought to have arisen from an ancient gene duplication event: Rad51, which is expressed in both mitosis and meiosis; and Dmc1, which is only expressed in meiosis. To explore the evolutionary relationship between these recombinases, here, we present high-resolution CryoEM structures of Rad51 filaments and Dmc1 filaments bound to ssDNA, which reveal a pair of stacked interfacial aromatic amino acid residues that are nearly universally conserved in Rad51 but are absent from Dmc1. We use a combination of bioinformatics, genetic analysis of natural sequence variation, and deep mutational analysis to probe the functionally tolerated sequence space for these stacked aromatic residues.

The separation pin distinguishes the pro- and anti-recombinogenic functions of Saccharomyces cerevisiae Srs2.

Srs2 is an Sf1a helicase that helps maintain genome stability in Saccharomyces cerevisiae through its ability to regulate homologous recombination. Srs2 downregulates HR by stripping Rad51 from single-stranded DNA, and Srs2 is also thought to promote synthesis-dependent strand annealing by unwinding D-loops. However, it has not been possible to evaluate the relative contributions of these two distinct activities to any aspect of recombination.

Checkpoint control in meiotic prophase: Idiosyncratic demands require unique characteristics.

Chromosomal transactions such as replication, recombination and segregation are monitored by cell cycle checkpoint cascades. These checkpoints ensure the proper execution of processes that are needed for faithful genome inheritance from one cell to the next, and across generations. In meiotic prophase, a specialized checkpoint monitors defining events of meiosis: programmed DNA break formation, followed by dedicated repair through recombination based on interhomolog (IH) crossovers.

Homeostatic Control of Meiotic Prophase Checkpoint Function by Pch2 and Hop1.

Checkpoint cascades link cell cycle progression with essential chromosomal processes. During meiotic prophase, recombination and chromosome synapsis are monitored by what are considered distinct checkpoints. In budding yeast, cells that lack the AAA+ ATPase Pch2 show an impaired cell cycle arrest in response to synapsis defects.

The small GTPase Arl8b regulates assembly of the mammalian HOPS complex on lysosomes.

The homotypic fusion and protein sorting (HOPS) complex is a multi-subunit complex conserved from yeast to mammals that regulates late endosome and lysosome fusion. However, little is known about how the HOPS complex is recruited to lysosomes in mammalian cells. Here, we report that the small GTPase Arl8b, but not Rab7 (also known as RAB7A), is essential for membrane localization of the human (h)Vps41 subunit of the HOPS complex.