RESUMO
G-quadruplexes (G4s) form throughout the genome and influence important cellular processes. Their deregulation can challenge DNA replication fork progression and threaten genome stability. Here, we demonstrate an unexpected role for the double-stranded DNA (dsDNA) translocase helicase-like transcription factor (HLTF) in responding to G4s. We show that HLTF, which is enriched at G4s in the human genome, can directly unfold G4s in vitro and uses this ATP-dependent translocase function to suppress G4 accumulation throughout the cell cycle. Additionally, MSH2 (a component of MutS heterodimers that bind G4s) and HLTF act synergistically to suppress G4 accumulation, restrict alternative lengthening of telomeres, and promote resistance to G4-stabilizing drugs. In a discrete but complementary role, HLTF restrains DNA synthesis when G4s are stabilized by suppressing primase-polymerase (PrimPol)-dependent repriming. Together, the distinct roles of HLTF in the G4 response prevent DNA damage and potentially mutagenic replication to safeguard genome stability.
RESUMO
Maintenance of DNA integrity is essential to all forms of life. DNA damage generated by reaction with genotoxic chemicals results in deleterious mutations, genome instability, and cell death. Pathogenic bacteria encounter several genotoxic agents during infection. In keeping with this, the loss of DNA repair networks results in virulence attenuation in several bacterial species. Interstrand DNA crosslinks (ICLs) are a type of DNA lesion formed by covalent linkage of opposing DNA strands and are particularly toxic as they interfere with replication and transcription. Bacteria have evolved specialized DNA glycosylases that unhook ICLs, thereby initiating their repair. In this study, we describe AlkX, a DNA glycosylase encoded by the multidrug resistant pathogen Acinetobacter baumannii. AlkX exhibits ICL unhooking activity similar to that of its Escherichia coli homolog YcaQ. Interrogation of the in vivo role of AlkX revealed that its loss sensitizes cells to DNA crosslinking and impairs A. baumannii colonization of the lungs and dissemination to distal tissues during pneumonia. These results suggest that AlkX participates in A. baumannii pathogenesis and protects the bacterium from stress conditions encountered in vivo. Consistent with this, we found that acidic pH, an environment encountered during host colonization, results in A. baumannii DNA damage and that alkX is induced by, and contributes to, defense against acidic conditions. Collectively, these studies reveal functions for a recently described class of proteins encoded in a broad range of pathogenic bacterial species.
Assuntos
Acinetobacter baumannii , Dano ao DNA , DNA Glicosilases , Acinetobacter baumannii/patogenicidade , Acinetobacter baumannii/genética , Acinetobacter baumannii/enzimologia , Acinetobacter baumannii/metabolismo , DNA Glicosilases/metabolismo , DNA Glicosilases/genética , Reparo do DNA , Infecções por Acinetobacter/microbiologia , Infecções por Acinetobacter/patologia , Proteínas de Bactérias/metabolismo , Proteínas de Bactérias/genética , Animais , Camundongos , DNA Bacteriano/genética , DNA Bacteriano/metabolismo , Virulência , Escherichia coli/genética , Escherichia coli/metabolismoRESUMO
Endonuclease VIII-like 3 (NEIL3) is a versatile DNA glycosylase that repairs a diverse array of chemical modifications to DNA. Unlike other glycosylases, NEIL3 has a preference for lesions within single-strand DNA and at single/double-strand DNA junctions. Beyond its canonical role in base excision repair of oxidized DNA, NEIL3 initiates replication-dependent interstrand DNA crosslink repair as an alternative to the Fanconi Anemia pathway. This review outlines our current understanding of NEIL3's biological functions, role in disease, and three-dimensional structure as it pertains to substrate specificity and catalytic mechanism.
Assuntos
DNA Glicosilases , Reparo do DNA , Humanos , DNA Glicosilases/metabolismo , DNA Glicosilases/química , Especificidade por Substrato , DNA/metabolismo , Dano ao DNA , Animais , Replicação do DNA , N-Glicosil HidrolasesRESUMO
The mechanism by which polymerase α-primase (polα-primase) synthesizes chimeric RNA-DNA primers of defined length and composition, necessary for replication fidelity and genome stability, is unknown. Here, we report cryo-EM structures of Xenopus laevis polα-primase in complex with primed templates representing various stages of DNA synthesis. Our data show how interaction of the primase regulatory subunit with the primer 5' end facilitates handoff of the primer to polα and increases polα processivity, thereby regulating both RNA and DNA composition. The structures detail how flexibility within the heterotetramer enables synthesis across two active sites and provide evidence that termination of DNA synthesis is facilitated by reduction of polα and primase affinities for the varied conformations along the chimeric primer-template duplex. Together, these findings elucidate a critical catalytic step in replication initiation and provide a comprehensive model for primer synthesis by polα-primase.
Assuntos
Microscopia Crioeletrônica , DNA Polimerase I , DNA Primase , Replicação do DNA , Modelos Moleculares , Xenopus laevis , DNA Primase/química , DNA Primase/metabolismo , DNA Primase/genética , DNA Polimerase I/metabolismo , DNA Polimerase I/química , Animais , Domínio Catalítico , DNA/metabolismo , DNA/química , DNA/biossíntese , Primers do DNA/metabolismo , Primers do DNA/genética , RNA/metabolismo , RNA/química , Conformação ProteicaRESUMO
Entre janeiro de 1985 e dezembro de 1993 foram tratadas seis meninas com prolapso de mucosa uretral através da exérese cirúrgica de prolapso e ampliaçäo do meato uretral externo. A patologia predominou em criança de pele negra e classe social baixa. Os principais sintomas e sinais foram sangramento vaginal e tumores a nível do meato uretral externo. Os resultados, com seguimento de um mês, três meses, nove meses, 12 meses e 24 meses foram excelentes