RNA:DNA hybrids from Okazaki fragments contribute to establish the Ku-mediated barrier to replication-fork degradation.

Nonhomologous end-joining (NHEJ) factors act in replication-fork protection, restart, and repair. Here, we identified a mechanism related to RNA:DNA hybrids to establish the NHEJ factor Ku-mediated barrier to nascent strand degradation in fission yeast. RNase H activities promote nascent strand degradation and replication restart, with a prominent role of RNase H2 in processing RNA:DNA hybrids to overcome the Ku barrier to nascent strand degradation. RNase H2 cooperates with the MRN-Ctp1 axis to sustain cell resistance to replication stress in a Ku-dependent manner. Mechanistically, the need of RNaseH2 in nascent strand degradation requires the primase activity that allows establishing the Ku barrier to Exo1, whereas impairing Okazaki fragment maturation reinforces the Ku barrier. Finally, replication stress induces Ku foci in a primase-dependent manner and favors Ku binding to RNA:DNA hybrids. We propose a function for the RNA:DNA hybrid originating from Okazaki fragments in controlling the Ku barrier specifying nuclease requirement to engage fork resection.

Non-homologous end-joining at challenged replication forks: an RNA connection?

Defective DNA replication, known as 'replication stress', is a source of DNA damage, a hallmark of numerous human diseases, including cancer, developmental defect, neurological disorders, and premature aging. Recent work indicates that non-homologous end-joining (NHEJ) is unexpectedly active during DNA replication to repair replication-born DNA lesions and to safeguard replication fork integrity. However, erroneous NHEJ events are deleterious to genome stability. RNAs are novel regulators of NHEJ activity through their ability to modulate the assembly of repair complexes in trans. At DNA damage sites, RNAs and DNA-embedded ribonucleotides modulate repair efficiency and fidelity. We discuss here how RNAs and associated proteins, including RNA binding proteins, may regulate NHEJ to sustain genome stability during DNA replication.

RIOK2 phosphorylation by RSK promotes synthesis of the human small ribosomal subunit.

Ribosome biogenesis lies at the nexus of various signaling pathways coordinating protein synthesis with cell growth and proliferation. This process is regulated by well-described transcriptional mechanisms, but a growing body of evidence indicates that other levels of regulation exist. Here we show that the Ras/mitogen-activated protein kinase (MAPK) pathway stimulates post-transcriptional stages of human ribosome synthesis. We identify RIOK2, a pre-40S particle assembly factor, as a new target of the MAPK-activated kinase RSK. RIOK2 phosphorylation by RSK stimulates cytoplasmic maturation of late pre-40S particles, which is required for optimal protein synthesis and cell proliferation. RIOK2 phosphorylation facilitates its release from pre-40S particles and its nuclear re-import, prior to completion of small ribosomal subunits. Our results bring a detailed mechanistic link between the Ras/MAPK pathway and the maturation of human pre-40S particles, which opens a hitherto poorly explored area of ribosome biogenesis.

Intragenic meiotic recombination in Schizosaccharomyces pombe is sensitive to environmental temperature changes.

Changes in environmental temperature influence cellular processes and their dynamics, and thus affect the life cycle of organisms that are unable to control their cell/body temperature. Meiotic recombination is the cellular process essential for producing healthy haploid gametes by providing physical links (chiasmata) between homologous chromosomes to guide their accurate segregation. Additionally, meiotic recombination-initiated by programmed DNA double-strand breaks (DSBs)-can generate genetic diversity and, therefore, is a driving force of evolution. Environmental temperature influencing meiotic recombination outcome thus may be a crucial determinant of reproductive success and genetic diversity. Indeed, meiotic recombination frequency in fungi, plants and invertebrates changes with temperature. In most organisms, these temperature-induced changes in meiotic recombination seem to be mediated through the meiosis-specific chromosome axis organization, the synaptonemal complex in particular. The fission yeast Schizosaccharomyces pombe does not possess a synaptonemal complex. Thus, we tested how environmental temperature modulates meiotic recombination frequency in the absence of a fully-fledged synaptonemal complex. We show that intragenic recombination (gene conversion) positively correlates with temperature within a certain range, especially at meiotic recombination hotspots. In contrast, crossover recombination, which manifests itself as chiasmata, is less affected. Based on our observations, we suggest that, in addition to changes in DSB frequency, DSB processing could be another temperature-sensitive step causing temperature-induced recombination rate alterations.