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1.
EMBO Rep ; 25(2): 725-744, 2024 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-38177923

RESUMO

Viral infection often trigger an ATM serine/threonine kinase (ATM)-dependent DNA damage response in host cells that suppresses viral replication. Viruses evolved different strategies to counteract this antiviral surveillance system. Here, we report that human herpesvirus 6B (HHV-6B) infection causes genomic instability by suppressing ATM signaling in host cells. Expression of immediate-early protein 1 (IE1) phenocopies this phenotype and blocks homology-directed double-strand break repair. Mechanistically, IE1 interacts with NBS1, and inhibits ATM signaling through two distinct domains. HHV-6B seems to efficiently inhibit ATM signaling as further depletion of either NBS1 or ATM do not significantly boost viral replication in infected cells. Interestingly, viral integration of HHV-6B into the host's telomeres is not strictly dependent on NBS1, challenging current models where integration occurs through homology-directed repair. Given that spontaneous IE1 expression has been detected in cells of subjects with inherited chromosomally-integrated form of HHV-6B (iciHHV-6B), a condition associated with several health conditions, our results raise the possibility of a link between genomic instability and the development of iciHHV-6-associated diseases.


Assuntos
Herpesvirus Humano 6 , Proteínas Imediatamente Precoces , Infecções por Roseolovirus , Humanos , Herpesvirus Humano 6/genética , Herpesvirus Humano 6/metabolismo , Infecções por Roseolovirus/genética , Proteínas Imediatamente Precoces/genética , Proteínas Imediatamente Precoces/metabolismo , Integração Viral , Instabilidade Genômica , Proteínas Mutadas de Ataxia Telangiectasia/genética , Proteínas Mutadas de Ataxia Telangiectasia/metabolismo
2.
Proc Natl Acad Sci U S A ; 116(39): 19552-19562, 2019 09 24.
Artigo em Inglês | MEDLINE | ID: mdl-31501315

RESUMO

High-risk human papillomaviruses (HR-HPVs) promote cervical cancer as well as a subset of anogenital and head and neck cancers. Due to their limited coding capacity, HPVs hijack the host cell's DNA replication and repair machineries to replicate their own genomes. How this host-pathogen interaction contributes to genomic instability is unknown. Here, we report that HPV-infected cancer cells express high levels of RNF168, an E3 ubiquitin ligase that is critical for proper DNA repair following DNA double-strand breaks, and accumulate high numbers of 53BP1 nuclear bodies, a marker of genomic instability induced by replication stress. We describe a mechanism by which HPV E7 subverts the function of RNF168 at DNA double-strand breaks, providing a rationale for increased homology-directed recombination in E6/E7-expressing cervical cancer cells. By targeting a new regulatory domain of RNF168, E7 binds directly to the E3 ligase without affecting its enzymatic activity. As RNF168 knockdown impairs viral genome amplification in differentiated keratinocytes, we propose that E7 hijacks the E3 ligase to promote the viral replicative cycle. This study reveals a mechanism by which tumor viruses reshape the cellular response to DNA damage by manipulating RNF168-dependent ubiquitin signaling. Importantly, our findings reveal a pathway by which HPV may promote the genomic instability that drives oncogenesis.


Assuntos
Quebras de DNA de Cadeia Dupla , Papillomaviridae/metabolismo , Proteínas E7 de Papillomavirus/metabolismo , Ubiquitina-Proteína Ligases/metabolismo , Linhagem Celular Tumoral , Reparo do DNA , Feminino , Instabilidade Genômica , Recombinação Homóloga , Interações Hospedeiro-Patógeno , Humanos , Proteínas E7 de Papillomavirus/genética , Infecções por Papillomavirus/genética , Infecções por Papillomavirus/virologia , Transdução de Sinais , Proteína 1 de Ligação à Proteína Supressora de Tumor p53/metabolismo , Ubiquitina/genética , Ubiquitina-Proteína Ligases/genética , Neoplasias do Colo do Útero/virologia
3.
Science ; 385(6711): eadl5816, 2024 Aug 23.
Artigo em Inglês | MEDLINE | ID: mdl-39088653

RESUMO

The human nucleosome acetyltransferase of histone H4 (NuA4)/Tat-interactive protein, 60 kilodalton (TIP60) coactivator complex, a fusion of the yeast switch/sucrose nonfermentable related 1 (SWR1) and NuA4 complexes, both incorporates the histone variant H2A.Z into nucleosomes and acetylates histones H4, H2A, and H2A.Z to regulate gene expression and maintain genome stability. Our cryo-electron microscopy studies show that, within the NuA4/TIP60 complex, the E1A binding protein P400 (EP400) subunit serves as a scaffold holding the different functional modules in specific positions, creating a distinct arrangement of the actin-related protein (ARP) module. EP400 interacts with the transformation/transcription domain-associated protein (TRRAP) subunit by using a footprint that overlaps with that of the Spt-Ada-Gcn5 acetyltransferase (SAGA) complex, preventing the formation of a hybrid complex. Loss of the TRRAP subunit leads to mislocalization of NuA4/TIP60, resulting in the redistribution of H2A.Z and its acetylation across the genome, emphasizing the dual functionality of NuA4/TIP60 as a single macromolecular assembly.


Assuntos
Montagem e Desmontagem da Cromatina , Lisina Acetiltransferase 5 , Humanos , Acetilação , Proteínas Adaptadoras de Transdução de Sinal , Microscopia Crioeletrônica , Proteínas de Ligação a DNA/química , Histonas/química , Lisina Acetiltransferase 5/química , Proteínas Nucleares/química , Nucleossomos/química , Nucleossomos/ultraestrutura , Domínios Proteicos , Fatores de Transcrição/química
4.
Life Sci Alliance ; 6(6)2023 06.
Artigo em Inglês | MEDLINE | ID: mdl-36882285

RESUMO

Spinal muscular atrophy is the leading genetic cause of infant mortality and results from depleted levels of functional survival of motor neuron (SMN) protein by either deletion or mutation of the SMN1 gene. SMN is characterized by a central TUDOR domain, which mediates the association of SMN with arginine methylated (Rme) partners, such as coilin, fibrillarin, and RNA pol II (RNA polymerase II). Herein, we biochemically demonstrate that SMN also associates with histone H3 monomethylated on lysine 79 (H3K79me1), defining SMN as not only the first protein known to associate with the H3K79me1 histone modification but also the first histone mark reader to recognize both methylated arginine and lysine residues. Mutational analyzes provide evidence that SMNTUDOR associates with H3 via an aromatic cage. Importantly, most SMNTUDOR mutants found in spinal muscular atrophy patients fail to associate with H3K79me1.


Assuntos
Código das Histonas , Atrofia Muscular Espinal , Proteína 1 de Sobrevivência do Neurônio Motor , Humanos , Lactente , Arginina , Lisina , Atrofia Muscular Espinal/genética , RNA Polimerase II , Fatores de Transcrição , Proteína 1 de Sobrevivência do Neurônio Motor/genética
5.
Nat Commun ; 14(1): 697, 2023 02 08.
Artigo em Inglês | MEDLINE | ID: mdl-36754959

RESUMO

Human acetyltransferases MOZ and MORF are implicated in chromosomal translocations associated with aggressive leukemias. Oncogenic translocations involve the far amino terminus of MOZ/MORF, the function of which remains unclear. Here, we identified and characterized two structured winged helix (WH) domains, WH1 and WH2, in MORF and MOZ. WHs bind DNA in a cooperative manner, with WH1 specifically recognizing unmethylated CpG sequences. Structural and genomic analyses show that the DNA binding function of WHs targets MORF/MOZ to gene promoters, stimulating transcription and H3K23 acetylation, and WH1 recruits oncogenic fusions to HOXA genes that trigger leukemogenesis. Cryo-EM, NMR, mass spectrometry and mutagenesis studies provide mechanistic insight into the DNA-binding mechanism, which includes the association of WH1 with the CpG-containing linker DNA and binding of WH2 to the dyad of the nucleosome. The discovery of WHs in MORF and MOZ and their DNA binding functions could open an avenue in developing therapeutics to treat diseases associated with aberrant MOZ/MORF acetyltransferase activities.


Assuntos
Acetiltransferases , Histona Acetiltransferases , Leucemia , Humanos , Acetilação , Acetiltransferases/metabolismo , Ilhas de CpG/genética , Histona Acetiltransferases/metabolismo , Leucemia/genética , Translocação Genética
6.
Cell Rep ; 39(11): 110947, 2022 06 14.
Artigo em Inglês | MEDLINE | ID: mdl-35705031

RESUMO

A recurrent chromosomal translocation found in acute myeloid leukemia leads to an in-frame fusion of the transcription repressor ZMYND11 to MBTD1, a subunit of the NuA4/TIP60 histone acetyltransferase complex. To understand the abnormal molecular events that ZMYND11-MBTD1 expression can create, we perform a biochemical and functional characterization comparison to each individual fusion partner. ZMYND11-MBTD1 is stably incorporated into the endogenous NuA4/TIP60 complex, leading to its mislocalization on the body of genes normally bound by ZMYND11. This can be correlated to increased chromatin acetylation and altered gene transcription, most notably on the MYC oncogene, and alternative splicing. Importantly, ZMYND11-MBTD1 expression favors Myc-driven pluripotency during embryonic stem cell differentiation and self-renewal of hematopoietic stem/progenitor cells. Altogether, these results indicate that the ZMYND11-MBTD1 fusion functions primarily by mistargeting the NuA4/TIP60 complex to the body of genes, altering normal transcription of specific genes, likely driving oncogenesis in part through the Myc regulatory network.


Assuntos
Cromatina , Histona Acetiltransferases , Proteínas de Fusão Oncogênica , Fases de Leitura Aberta , Acetilação , Proteínas de Ciclo Celular/metabolismo , Proteínas Cromossômicas não Histona/metabolismo , Proteínas Correpressoras/metabolismo , Proteínas de Ligação a DNA/metabolismo , Histona Acetiltransferases/genética , Histona Acetiltransferases/metabolismo , Humanos , Lisina Acetiltransferase 5/genética , Lisina Acetiltransferase 5/metabolismo , Proteínas de Fusão Oncogênica/genética , Proteínas de Fusão Oncogênica/metabolismo , Fases de Leitura Aberta/genética , Translocação Genética
7.
Front Cell Dev Biol ; 9: 729338, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34604228

RESUMO

The modification of histones-the structural components of chromatin-is a central topic in research efforts to understand the mechanisms regulating genome expression and stability. These modifications frequently occur through associations with multisubunit complexes, which contain active enzymes and additional components that orient their specificity and read the histone modifications that comprise epigenetic signatures. To understand the functions of these modifications it is critical to study the enzymes and substrates involved in their native contexts. Here, we describe experimental approaches to purify native chromatin modifiers complexes from mammalian cells and to produce recombinant nucleosomes that are used as substrates to determine the activity of the complex. In addition, we present a novel approach, similar to the yeast anchor-away system, to study the functions of essential chromatin modifiers by quickly inducing their depletion from the nucleus. The step-by-step protocols included will help standardize these approaches in the research community, enabling convincing conclusions about the specificities and functions of these crucial regulators of the eukaryotic genome.

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