Chd7 is indispensable for mammalian brain development through activation of a neuronal differentiation programme

Mutations in chromatin modifier genes are frequently associated with neurodevelopmental diseases. We herein demonstrate that the chromodomain helicase DNA-binding protein 7 (Chd7), frequently associated with CHARGE syndrome, is indispensable for normal cerebellar development. Genetic inactivation of Chd7 in cerebellar granule neuron progenitors leads to cerebellar hypoplasia in mice, due to the impairment of granule neuron differentiation, induction of apoptosis and abnormal localization of Purkinje cells, which closely recapitulates known clinical features in the cerebella of CHARGE patients. Combinatory molecular analyses reveal that Chd7 is required for the maintenance of open chromatin and thus activation of genes essential for granule neuron differentiation. We further demonstrate that both Chd7 and Top2b are necessary for the transcription of a set of long neuronal genes in cerebellar granule neurons. Altogether, our comprehensive analyses reveal a mechanism with chromatin remodellers governing brain development via controlling a core transcriptional programme for cell-specific differentiation.Spanish Government SAF2010-21017, SAF2013-47343-P, SAF2014-55532-RAndalusian Regional Government P11-CVI-794

ATM specifically mediates repair of double-strand breaks with blocked DNA ends

Ataxia telangiectasia is caused by mutations in ATM and represents a paradigm for cancer predisposition and neurodegenerative syndromes linked to deficiencies in the DNA-damage response. The role of ATM as a key regulator of signalling following DNA double-strand breaks (DSBs) has been dissected in extraordinary detail, but the impact of this process on DSB repair still remains controversial. Here we develop novel genetic and molecular tools to modify the structure of DSB ends and demonstrate that ATM is indeed required for efficient and accurate DSB repair, preventing cell death and genome instability, but exclusively when the ends are irreversibly blocked. We therefore identify the nature of ATM involvement in DSB repair, presenting blocked DNA ends as a possible pathogenic trigger of ataxia telangiectasia and related disorders

ATM specifically mediates repair of double-strand breaks with blocked DNA ends

Ataxia telangiectasia is caused by mutations in ATM and represents a paradigm for cancer predisposition and neurodegenerative syndromes linked to deficiencies in the DNA-damage response. The role of ATM as a key regulator of signalling following DNA double-strand breaks (DSBs) has been dissected in extraordinary detail, but the impact of this process on DSB repair still remains controversial. Here we develop novel genetic and molecular tools to modify the structure of DSB ends and demonstrate that ATM is indeed required for efficient and accurate DSB repair, preventing cell death and genome instability, but exclusively when the ends are irreversibly blocked. We therefore identify the nature of ATM involvement in DSB repair, presenting blocked DNA ends as a possible pathogenic trigger of ataxia telangiectasia and related disorders

Regulation of TDP2 functions by SUMO interactions

Topoisomerase 2 (TOP2) performs a vital enzymatic activity, solving DNA topological problems in fundamental metabolic processes. The enzyme is able to untangle DNA by passing an intact helix through a transient double-strand break (DSB). The intermediate of its catalytic cycle, TOP2 covalently bound to DNA, is usually short lived, and is known as the TOP2 cleavage complex (TOP2cc). Tyrosyl DNA phosphodiesterase 2 (TDP2) is a protein involved in the removal of proteasome degraded TOP2cc, thus im-portant for the efficient repair of TOP2-induced DNA DSBs. Consequently, TDP2-deficient cells are sensitive to the TOP2 poison etoposide. Sumoylation is an important modification involved in the DNA damage response (DDR). While TOP2 sumoylation is important in several cellular processes, it is not known whether TOP2 is sumoylated in the context of the cleavage complex and the possible physiological relevance. In this work, Here, we describe for the first time SUMO1 and SUMO2/3 modification of TOP2 in the context of the cleavage complex. We imply a role for the novel TDP2 split-SIM in the preferential removal of SUMO modified TOP2cc. In order to identify pos-sible interactors involved in this process, we performed a BioID screening and identify the novel SUMO E3 ligase ZNF451, along with potential TDP2 involvement in novel processes. Through a BioID screening, we identify possible new TDP2 interactors and TDP2 involvement in novel processes. We discover a novel pathway for the repair of TOP2cc independent of known proteasomal pathways, and imply a role of the novel SUMO E3 ligase ZNF451. We suggest a role for the novel TDP2 split-SIM in the pref-erential removal of SUMO modified TOP2cc. Additionally, we find evidence for a TDP2 and proteasome independent role for ZNF451 in the repair of TOP2 double-strand breaks (DSBs).Peer Reviewe

Regulation of the newly discovered human Tdp2 enzyme by SUMO and Ubiquitin

Póster presentado al 22nd IUBMB & 37th FEBS Congress: From Single Molecules to Systems Biology, celebrado en Sevilla (España) del 4 al 9 de septiembre de 2012.Peer Reviewe

ATM specifically mediates repair of double-strand breaks with blocked DNA ends

Ataxia telangiectasia is caused by mutations in ATM and represents a paradigm for cancer predisposition and neurodegenerative syndromes linked to deficiencies in the DNA-damage response. The role of ATM as a key regulator of signalling following DNA double-strand breaks (DSBs) has been dissected in extraordinary detail, but the impact of this process on DSB repair still remains controversial. Here we develop novel genetic and molecular tools to modify the structure of DSB ends and demonstrate that ATM is indeed required for efficient and accurate DSB repair, preventing cell death and genome instability, but exclusively when the ends are irreversibly blocked. We therefore identify the nature of ATM involvement in DSB repair, presenting blocked DNA ends as a possible pathogenic trigger of ataxia telangiectasia and related disordersWork in F.C.-L. laboratory is funded with grants from the Spanish Government (SAF2010-21017 and BFU2010-11042-E, Ministerio de Ciencia e Innovación), the regional Andalusian Government (CVI-7948) and the European Union (PERG07-2010-268466) and with the following fellowships: Formación Personal Investigador (BES-2011-047351, Ministerio de Ciencia e Innovación) for A.A.-Q., Beca Predoctoral AEFAT (Asociación Española Familia Ataxia Telangiectasia) for A.S.-B., Personal Investigador en Formación (Universidad de Sevilla) for J.A.L. and Ramón y Cajal (RYC-2009-03928, Ministerio de Ciencia e Innovación) for F.C.-L. L.M.E. is supported by the Miguel Servet program, (Instituto Carlos III) and the Spanish Government grant (BFU2011-25734, Ministerio de Ciencia e Innovación)Peer Reviewe

Chd7 is indispensable for mammalian brain development through activation of a neuronal differentiation programme

Feng, Weijun et al.Mutations in chromatin modifier genes are frequently associated with neurodevelopmental diseases. We herein demonstrate that the chromodomain helicase DNA-binding protein 7 (Chd7), frequently associated with CHARGE syndrome, is indispensable for normal cerebellar development. Genetic inactivation of Chd7 in cerebellar granule neuron progenitors leads to cerebellar hypoplasia in mice, due to the impairment of granule neuron differentiation, induction of apoptosis and abnormal localization of Purkinje cells, which closely recapitulates known clinical features in the cerebella of CHARGE patients. Combinatory molecular analyses reveal that Chd7 is required for the maintenance of open chromatin and thus activation of genes essential for granule neuron differentiation. We further demonstrate that both Chd7 and Top2b are necessary for the transcription of a set of long neuronal genes in cerebellar granule neurons. Altogether, our comprehensive analyses reveal a mechanism with chromatin remodellers governing brain development via controlling a core transcriptional programme for cell-specific differentiation.This work was supported by the Helmholtz Association (VH-NG-702), the Deutsche Forschungsgemeinschaft (KA 4472/1-1 for D.K., LI 2140/1-1 for H.-K.L.), the Deutsche Krebshilfe (110226 to H.-K.L.), the ERC (European Research Council) consolidator grant (647055) (to H.-K.L.), the Helmholtz Alliance ‘Preclinical Comprehensive Cancer Center' Grant HA-305 (to J.G., P.L. S.M.P. and H.-K.L.), the DKFZ Intramural Grant (to W.F. and D.K.), the Spanish Government (SAF2010-21017, SAF2013-47343-P, SAF2014-55532-R and FEDER funds, to F.C.-L.), the Andalusian Regional Government (P11-CVI-7948 and FEDER funds, to F.C.-L.), the European Research Council (ERC-CoG-2014-647359, to F.C.-L.) and Predoctoral Studentships from the University of Seville to J.A.L. (PIF-2011). CABIMER is supported by the Andalusian Regional Government (Junta de Andalucía).Peer Reviewe

ZATT (ZNF451)–mediated resolution of topoisomerase 2 DNA-protein cross-links

Schellenberg, Matthew J. et al.Topoisomerase 2 (TOP2) DNA transactions proceed via formation of the TOP2 cleavage complex (TOP2cc), a covalent enzyme-DNA reaction intermediate that is vulnerable to trapping by potent anticancer TOP2 drugs. How genotoxic TOP2 DNA-protein cross-links are resolved is unclear. We found that the SUMO (small ubiquitin-related modifier) ligase ZATT (ZNF451) is a multifunctional DNA repair factor that controls cellular responses to TOP2 damage. ZATT binding to TOP2cc facilitates a proteasome-independent tyrosyl-DNA phosphodiesterase 2 (TDP2) hydrolase activity on stalled TOP2cc. The ZATT SUMO ligase activity further promotes TDP2 interactions with SUMOylated TOP2, regulating efficient TDP2 recruitment through a “split-SIM” SUMO2 engagement platform. These findings uncover a ZATT-TDP2–catalyzed and SUMO2-modulated pathway for direct resolution of TOP2cc.Supported by NIH Intramural Research Program grants 1Z01ES102765 (R.S.W.), 1ZIAES050111-26 (R.E.L.), and ZES102488-09 (J.G.W.); Spanish and Andalusian Government grants SAF2010-21017, SAF2013-47343-P, SAF2014-55532-R, CVI-7948, and FEDER funds (F.C.-L.) and BES-2015-071672 (A.H.-R.); European Research Council grant ERC-CoG-2014-647359 (F.C.-L.); and University of Seville grant PIF-2011 (J.A.L.). The Advanced Photon Source SERCAT beamline is supported by the U.S. Department of Energy, Office of Basic Energy Sciences (DOE OBES) grant W-31-109-Eng-38. The Advanced Light Source is operated by Lawrence Berkeley National Laboratory on behalf of DOE OBES and the IDAT program, supported by the DOE Office of Biological and Environmental Research and NIH project MINOS (R01GM105404). CABIMER is supported by the Andalusian Government.Peer Reviewe

Topoisomerase IIα represses transcription by enforcing promoter-proximal pausing

Accumulation of topological stress in the form of DNA supercoiling is inherent to the advance of RNA polymerase II (Pol II) and needs to be resolved by DNA topoisomerases to sustain productive transcriptional elongation. Topoisomerases are therefore considered positive facilitators of transcription. Here, we show that, in contrast to this general assumption, human topoisomerase IIα (TOP2A) activity at promoters represses transcription of immediate early genes such as c-FOS, maintaining them under basal repressed conditions. Thus, TOP2A inhibition creates a particular topological context that results in rapid release from promoter-proximal pausing and transcriptional upregulation, which mimics the typical bursting behavior of these genes in response to physiological stimulus. We therefore describe the control of promoter-proximal pausing by TOP2A as a layer for the regulation of gene expression, which can act as a molecular switch to rapidly activate transcription, possibly by regulating the accumulation of DNA supercoiling at promoter regions.This work was funded with grants from the Spanish and Andalusian governments (SAF2017-89619-R, CVI-7948, and European Regional Development Fund) and the European Research Council (ERC-CoG-2014-647359), and with individual fellowships for A.H.-R. (Contratos para la Formación de Doctores, BES-2015-071672, and Ministerio de Economía y Competitividad); S.J.-G. (Ramó n y Cajal, RYC-2015-17246, and Ministerio de Economía y Competitividad); J.T.-B. (Formación Profesorado Universitario, FPU15/03656, and Ministerio de Educación, Cultura y Deporte); and G.M.-Z. (AECC Postdoctoral Fellowships). CABIMER is supported by the Andalusian Government.Ye

ZATT (ZNF451)-mediated resolution of topoisomerase 2 DNA-protein cross-links.

Topoisomerase 2 (TOP2) DNA transactions proceed via formation of the TOP2 cleavage complex (TOP2cc), a covalent enzyme-DNA reaction intermediate that is vulnerable to trapping by potent anticancer TOP2 drugs. How genotoxic TOP2 DNA-protein cross-links are resolved is unclear. We found that the SUMO (small ubiquitin-related modifier) ligase ZATT (ZNF451) is a multifunctional DNA repair factor that controls cellular responses to TOP2 damage. ZATT binding to TOP2cc facilitates a proteasome-independent tyrosyl-DNA phosphodiesterase 2 (TDP2) hydrolase activity on stalled TOP2cc. The ZATT SUMO ligase activity further promotes TDP2 interactions with SUMOylated TOP2, regulating efficient TDP2 recruitment through a "split-SIM" SUMO2 engagement platform. These findings uncover a ZATT-TDP2-catalyzed and SUMO2-modulated pathway for direct resolution of TOP2cc