ATM Dependent Silencing Links Nucleolar Chromatin Reorganization to DNA Damage Recognition

SummaryResolution of DNA double-strand breaks (DSBs) is essential for the suppression of genome instability. DSB repair in transcriptionally active genomic regions represents a unique challenge that is associated with ataxia telangiectasia mutated (ATM) kinase-mediated transcriptional silencing. Despite emerging insights into the underlying mechanisms, how DSB silencing connects to DNA repair remains undefined. We observe that silencing within the rDNA depends on persistent DSBs. Non-homologous end-joining was the predominant mode of DSB repair allowing transcription to resume. ATM-dependent rDNA silencing in the presence of persistent DSBs led to the large-scale reorganization of nucleolar architecture, with movement of damaged chromatin to nucleolar cap regions. These findings identify ATM-dependent temporal and spatial control of DNA repair and provide insights into how communication between DSB signaling and ongoing transcription promotes genome integrity

Nuclear rupture at sites of high curvature compromises retention of DNA repair factors.

The nucleus is physically linked to the cytoskeleton, adhesions, and extracellular matrix-all of which sustain forces, but their relationships to DNA damage are obscure. We show that nuclear rupture with cytoplasmic mislocalization of multiple DNA repair factors correlates with high nuclear curvature imposed by an external probe or by cell attachment to either aligned collagen fibers or stiff matrix. Mislocalization is greatly enhanced by lamin A depletion, requires hours for nuclear reentry, and correlates with an increase in pan-nucleoplasmic foci of the DNA damage marker γH2AX. Excess DNA damage is rescued in ruptured nuclei by cooverexpression of multiple DNA repair factors as well as by soft matrix or inhibition of actomyosin tension. Increased contractility has the opposite effect, and stiff tumors with low lamin A indeed exhibit increased nuclear curvature, more frequent nuclear rupture, and excess DNA damage. Additional stresses likely play a role, but the data suggest high curvature promotes nuclear rupture, which compromises retention of DNA repair factors and favors sustained damage

Rescue of DNA damage after constricted migration reveals a mechano-regulated threshold for cell cycle.

Migration through 3D constrictions can cause nuclear rupture and mislocalization of nuclear proteins, but damage to DNA remains uncertain, as does any effect on cell cycle. Here, myosin II inhibition rescues rupture and partially rescues the DNA damage marker γH2AX, but an apparent block in cell cycle appears unaffected. Co-overexpression of multiple DNA repair factors or antioxidant inhibition of break formation also exert partial effects, independently of rupture. Combined treatments completely rescue cell cycle suppression by DNA damage, revealing a sigmoidal dependence of cell cycle on excess DNA damage. Migration through custom-etched pores yields the same damage threshold, with ∼4-µm pores causing intermediate levels of both damage and cell cycle suppression. High curvature imposed rapidly by pores or probes or else by small micronuclei consistently associates nuclear rupture with dilution of stiff lamin-B filaments, loss of repair factors, and entry from cytoplasm of chromatin-binding cGAS (cyclic GMP-AMP synthase). The cell cycle block caused by constricted migration is nonetheless reversible, with a potential for DNA misrepair and genome variation

Diets containing sea cucumber (Isostichopus badionotus) meals are hypocholesterolemic in young rats

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Case-control analysis of truncating mutations in DNA damage response genes connects TEX15 and FANCD2 with hereditary breast cancer susceptibility

Several known breast cancer susceptibility genes encode proteins involved in DNA damage response (DDR) and are characterized by rare loss-of-function mutations. However, these explain less than half of the familial cases. To identify novel susceptibility factors, 39 rare truncating mutations, identified in 189 Northern Finnish hereditary breast cancer patients in parallel sequencing of 796 DDR genes, were studied for disease association. Mutation screening was performed for Northern Finnish breast cancer cases (n = 578-1565) and controls (n = 337-1228). Mutations showing potential cancer association were analyzed in additional Finnish cohorts.c.7253dupT in TEX15, encoding a DDR factor important in meiosis, associated with hereditary breast cancer (p = 0.018) and likely represents a Northern Finnish founder mutation. A deleterious c.2715 + 1G > A mutation in the Fanconi anemia gene, FANCD2, was over two times more common in the combined Finnish hereditary cohort compared to controls. A deletion (c.640_644del5) in RNF168, causative for recessive RIDDLE syndrome, had high prevalence in majority of the analyzed cohorts, but did not associate with breast cancer. In conclusion, truncating variants in TEX15 and FANCD2 are potential breast cancer risk factors, warranting further investigations in other populations. Furthermore, high frequency of RNF168 c.640_644del5 indicates the need for its testing in Finnish patients with RIDDLE syndrome symptoms.Peer reviewe

Higher-Order Assembly of BRCC36-KIAA0157 Is Required for DUB Activity and Biological Function

BRCC36 is a Zn²⁺-dependent deubiquitinating enzyme (DUB) that hydrolyzes lysine-63-linked ubiquitin chains as part of distinct macromolecular complexes that participate in either interferon signaling or DNA-damage recognition. The MPN⁺ domain protein BRCC36 associates with pseudo DUB MPN⁻ proteins KIAA0157 or Abraxas, which are essential for BRCC36 enzymatic activity. To understand the basis for BRCC36 regulation, we have solved the structure of an active BRCC36-KIAA0157 heterodimer and an inactive BRCC36 homodimer. Structural and functional characterizations show how BRCC36 is switched to an active conformation by contacts with KIAA0157. Higher-order association of BRCC36 and KIAA0157 into a dimer of heterodimers (super dimers) was required for DUB activity and interaction with targeting proteins SHMT2 and RAP80. These data provide an explanation of how an inactive pseudo DUB allosterically activates a cognate DUB partner and implicates super dimerization as a new regulatory mechanism underlying BRCC36 DUB activity, subcellular localization, and biological function

Efficacy of a monovalent human-bovine (116E) rotavirus vaccine in Indian children in the second year of life

Rotavirus gastroenteritis is one of the leading causes of diarrhea in Indian children less than 2 years of age. The 116E rotavirus strain was developed as part of the Indo-US Vaccine Action Program and has undergone efficacy trials. This paper reports the efficacy and additional safety data in children up to 2 years of age. In a double-blind placebo controlled multicenter trial, 6799 infants aged 6-7 weeks were randomized to receive three doses of an oral human-bovine natural reassortant vaccine (116E) or placebo at ages 6, 10, and 14 weeks. The primary outcome was severe (≥11 on the Vesikari scale) rotavirus gastroenteritis. Efficacy outcomes and adverse events were ascertained through active surveillance. We randomly assigned 4532 and 2267 subjects to receive vaccine and placebo, respectively, with over 96% subjects receiving all three doses of the vaccine or placebo. The per protocol analyses included 4354 subjects in the vaccine and 2187 subjects in the placebo group. The overall incidence of severe RVGE per 100 person years was 1.3 in the vaccine group and 2.9 in the placebo recipients. Vaccine efficacy against severe rotavirus gastroenteritis in children up to 2 years of age was 55.1% (95% CI 39.9 to 66.4; p<0.0001); vaccine efficacy in the second year of life of 48.9% (95% CI 17.4 to 68.4; p=0.0056) was only marginally less than in the first year of life [56.3% (95% CI 36.7 to 69.9; p<0.0001)]. The number of infants needed to be immunized to prevent one episode of severe RVGE in the first 2 years of life was 40 (95% CI 28.0 to 63.0) and for RVGE of any severity, it was 21 (95% CI 16.0 to 32.0). Serious adverse events were observed at the same rates in the two groups. None of the eight intussusception events occurred within 30 days of a vaccine dose and all were reported only after the third dose. The sustained efficacy of the 116E in the second year of life is reassuring