+Open data
-Basic information
Entry | Database: PDB / ID: 6upl | ||||||
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Title | Structure of FACT_subnucleosome complex 2 | ||||||
Components |
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Keywords | TRANSCRIPTION/DNA / nucleosome assembly / nucleosome disassembly / transient / integrity / TRANSCRIPTION / replication / histone chaperone / TRANSCRIPTION-DNA complex | ||||||
Function / homology | Function and homology information FACT complex / regulation of chromatin organization / nucleosome disassembly / positive regulation of DNA-templated transcription, elongation / Pausing and recovery of Tat-mediated HIV elongation / Tat-mediated HIV elongation arrest and recovery / HIV elongation arrest and recovery / Pausing and recovery of HIV elongation / nucleosome binding / Tat-mediated elongation of the HIV-1 transcript ...FACT complex / regulation of chromatin organization / nucleosome disassembly / positive regulation of DNA-templated transcription, elongation / Pausing and recovery of Tat-mediated HIV elongation / Tat-mediated HIV elongation arrest and recovery / HIV elongation arrest and recovery / Pausing and recovery of HIV elongation / nucleosome binding / Tat-mediated elongation of the HIV-1 transcript / Formation of HIV-1 elongation complex containing HIV-1 Tat / negative regulation of megakaryocyte differentiation / Formation of HIV elongation complex in the absence of HIV Tat / protein localization to CENP-A containing chromatin / Chromatin modifying enzymes / Replacement of protamines by nucleosomes in the male pronucleus / CENP-A containing nucleosome / RNA Polymerase II Transcription Elongation / Formation of RNA Pol II elongation complex / Packaging Of Telomere Ends / epigenetic regulation of gene expression / Recognition and association of DNA glycosylase with site containing an affected purine / Cleavage of the damaged purine / Deposition of new CENPA-containing nucleosomes at the centromere / RNA Polymerase II Pre-transcription Events / Recognition and association of DNA glycosylase with site containing an affected pyrimidine / Cleavage of the damaged pyrimidine / Inhibition of DNA recombination at telomere / Meiotic synapsis / telomere organization / RNA Polymerase I Promoter Opening / Interleukin-7 signaling / SUMOylation of chromatin organization proteins / Assembly of the ORC complex at the origin of replication / DNA methylation / Condensation of Prophase Chromosomes / SIRT1 negatively regulates rRNA expression / HCMV Late Events / Chromatin modifications during the maternal to zygotic transition (MZT) / ERCC6 (CSB) and EHMT2 (G9a) positively regulate rRNA expression / innate immune response in mucosa / PRC2 methylates histones and DNA / Defective pyroptosis / transcription elongation by RNA polymerase II / HDACs deacetylate histones / TP53 Regulates Transcription of DNA Repair Genes / RNA Polymerase I Promoter Escape / Nonhomologous End-Joining (NHEJ) / Transcriptional regulation by small RNAs / Formation of the beta-catenin:TCF transactivating complex / RUNX1 regulates genes involved in megakaryocyte differentiation and platelet function / Activated PKN1 stimulates transcription of AR (androgen receptor) regulated genes KLK2 and KLK3 / NoRC negatively regulates rRNA expression / G2/M DNA damage checkpoint / B-WICH complex positively regulates rRNA expression / HDMs demethylate histones / DNA Damage/Telomere Stress Induced Senescence / Metalloprotease DUBs / PKMTs methylate histone lysines / RMTs methylate histone arginines / Meiotic recombination / Pre-NOTCH Transcription and Translation / nucleosome assembly / Activation of anterior HOX genes in hindbrain development during early embryogenesis / HCMV Early Events / structural constituent of chromatin / Transcriptional regulation of granulopoiesis / UCH proteinases / nucleosome / antimicrobial humoral immune response mediated by antimicrobial peptide / E3 ubiquitin ligases ubiquitinate target proteins / gene expression / Recruitment and ATM-mediated phosphorylation of repair and signaling proteins at DNA double strand breaks / RUNX1 regulates transcription of genes involved in differentiation of HSCs / chromatin organization / Factors involved in megakaryocyte development and platelet production / HATs acetylate histones / Processing of DNA double-strand break ends / antibacterial humoral response / histone binding / Senescence-Associated Secretory Phenotype (SASP) / Oxidative Stress Induced Senescence / Estrogen-dependent gene expression / Regulation of TP53 Activity through Phosphorylation / DNA replication / chromosome, telomeric region / transcription by RNA polymerase II / Ub-specific processing proteases / defense response to Gram-positive bacterium / cadherin binding / Amyloid fiber formation / protein heterodimerization activity / negative regulation of cell population proliferation / DNA repair / nucleolus / protein-containing complex / DNA binding / extracellular space / RNA binding / extracellular exosome Similarity search - Function | ||||||
Biological species | Homo sapiens (human) Escherichia coli (E. coli) | ||||||
Method | ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 7.4 Å | ||||||
Authors | Zhou, K. / Tan, Y.Z. / Wei, H. / Liu, Y. / Carragher, B. / Potter, C. / Luger, K. | ||||||
Funding support | United States, 1items
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Citation | Journal: Nature / Year: 2020 Title: FACT caught in the act of manipulating the nucleosome. Authors: Yang Liu / Keda Zhou / Naifu Zhang / Hui Wei / Yong Zi Tan / Zhening Zhang / Bridget Carragher / Clinton S Potter / Sheena D'Arcy / Karolin Luger / Abstract: The organization of genomic DNA into nucleosomes profoundly affects all DNA-related processes in eukaryotes. The histone chaperone known as 'facilitates chromatin transcription' (FACT) (consisting of ...The organization of genomic DNA into nucleosomes profoundly affects all DNA-related processes in eukaryotes. The histone chaperone known as 'facilitates chromatin transcription' (FACT) (consisting of subunits SPT16 and SSRP1) promotes both disassembly and reassembly of nucleosomes during gene transcription, DNA replication and DNA repair. However, the mechanism by which FACT causes these opposing outcomes is unknown. Here we report two cryo-electron-microscopic structures of human FACT in complex with partially assembled subnucleosomes, with supporting biochemical and hydrogen-deuterium exchange data. We find that FACT is engaged in extensive interactions with nucleosomal DNA and all histone variants. The large DNA-binding surface on FACT appears to be protected by the carboxy-terminal domains of both of its subunits, and this inhibition is released by interaction with H2A-H2B, allowing FACT-H2A-H2B to dock onto a complex containing DNA and histones H3 and H4 (ref. ). SPT16 binds nucleosomal DNA and tethers H2A-H2B through its carboxy-terminal domain by acting as a placeholder for DNA. SSRP1 also contributes to DNA binding, and can assume two conformations, depending on whether a second H2A-H2B dimer is present. Our data suggest a compelling mechanism for how FACT maintains chromatin integrity during polymerase passage, by facilitating removal of the H2A-H2B dimer, stabilizing intermediate subnucleosomal states and promoting nucleosome reassembly. Our findings reconcile discrepancies regarding the many roles of FACT and underscore the dynamic interactions between histone chaperones and nucleosomes. | ||||||
History |
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-Structure visualization
Movie |
Movie viewer |
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Structure viewer | Molecule: MolmilJmol/JSmol |
-Downloads & links
-Download
PDBx/mmCIF format | 6upl.cif.gz | 382.9 KB | Display | PDBx/mmCIF format |
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PDB format | pdb6upl.ent.gz | 294.2 KB | Display | PDB format |
PDBx/mmJSON format | 6upl.json.gz | Tree view | PDBx/mmJSON format | |
Others | Other downloads |
-Validation report
Arichive directory | https://data.pdbj.org/pub/pdb/validation_reports/up/6upl ftp://data.pdbj.org/pub/pdb/validation_reports/up/6upl | HTTPS FTP |
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-Related structure data
Related structure data | 20841MC 6upkC M: map data used to model this data C: citing same article (ref.) |
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Similar structure data | |
EM raw data | EMPIAR-10333 (Title: Single Particle Cryo-EM Reconstructions of Human FACT in Complex with Partially Assembled Sub-nucleosomes Data size: 2.2 TB Data #1: Unaligned multi-frame micrographs [micrographs - multiframe] Data #2: Aligned and dose-weighted micrographs [micrographs - single frame] Data #3: Final Particle Stacks for Class 1 and 2 with Euler Angles and Shifts [picked particles - single frame - processed]) |
-Links
-Assembly
Deposited unit |
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1 |
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-Components
-Protein , 4 types, 8 molecules AEBFCKDL
#1: Protein | Mass: 15437.167 Da / Num. of mol.: 2 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Homo sapiens (human) Gene: H3C1, H3FA, HIST1H3A, H3C2, H3FL, HIST1H3B, H3C3, H3FC HIST1H3C, H3C4, H3FB, HIST1H3D, H3C6, H3FD, HIST1H3E, H3C7, H3FI, HIST1H3F, H3C8, H3FH, HIST1H3G, H3C10, H3FK, HIST1H3H, H3C11, H3FF, ...Gene: H3C1, H3FA, HIST1H3A, H3C2, H3FL, HIST1H3B, H3C3, H3FC HIST1H3C, H3C4, H3FB, HIST1H3D, H3C6, H3FD, HIST1H3E, H3C7, H3FI, HIST1H3F, H3C8, H3FH, HIST1H3G, H3C10, H3FK, HIST1H3H, H3C11, H3FF, HIST1H3I, H3C12, H3FJ, HIST1H3J Production host: Escherichia coli 'BL21-Gold(DE3)pLysS AG' (bacteria) References: UniProt: P68431 #2: Protein | Mass: 11394.426 Da / Num. of mol.: 2 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Homo sapiens (human) Gene: HIST1H4A, H4/A, H4FA, HIST1H4B, H4/I, H4FI, HIST1H4C, H4/G, H4FG, HIST1H4D, H4/B, H4FB, HIST1H4E, H4/J, H4FJ, HIST1H4F, H4/C, H4FC, HIST1H4H, H4/H, H4FH, HIST1H4I, H4/M, H4FM, HIST1H4J, H4/E, ...Gene: HIST1H4A, H4/A, H4FA, HIST1H4B, H4/I, H4FI, HIST1H4C, H4/G, H4FG, HIST1H4D, H4/B, H4FB, HIST1H4E, H4/J, H4FJ, HIST1H4F, H4/C, H4FC, HIST1H4H, H4/H, H4FH, HIST1H4I, H4/M, H4FM, HIST1H4J, H4/E, H4FE, HIST1H4K, H4/D, H4FD, HIST1H4L, H4/K, H4FK, HIST2H4A, H4/N, H4F2, H4FN, HIST2H4, HIST2H4B, H4/O, H4FO, HIST4H4 Production host: Escherichia coli 'BL21-Gold(DE3)pLysS AG' (bacteria) References: UniProt: P62805 #3: Protein | Mass: 14135.523 Da / Num. of mol.: 2 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Homo sapiens (human) / Gene: HIST1H2AC, H2AFL Production host: Escherichia coli 'BL21-Gold(DE3)pLysS AG' (bacteria) References: UniProt: Q93077 #4: Protein | Mass: 13937.213 Da / Num. of mol.: 2 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Homo sapiens (human) Gene: HIST1H2BC, H2BFL, HIST1H2BE, H2BFH, HIST1H2BF, H2BFG, HIST1H2BG, H2BFA, HIST1H2BI, H2BFK Production host: Escherichia coli 'BL21-Gold(DE3)pLysS AG' (bacteria) References: UniProt: P62807 |
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-FACT complex subunit ... , 2 types, 2 molecules GH
#5: Protein | Mass: 109574.867 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Details: Amino acids 640-651 cannot be identified due to the resolution limitations. To show the connectivity of the electron density, poly-(UNK) was placed. This was also done for the end of this ...Details: Amino acids 640-651 cannot be identified due to the resolution limitations. To show the connectivity of the electron density, poly-(UNK) was placed. This was also done for the end of this chain (926-967). The full sequence for the experiment is: MHHHHHHAVTLDKDAYYRRVKRLYSNWRKGEDEYANVDAIVVSVGVDEEIVYAKSTALQTWLFGYELTDTIMVFCDDKII FMASKKKVEFLKQIANTKGNENANGAPAITLLIREKNESNKSSFDKMIEAIKESKNGKKIGVFSKDKFPGEFMKSWNDCL NKEGFDKIDISAVVAYTIAVKEDGELNLMKKAASITSEVFNKFFKERVMEIVDADEKVRHSKLAESVEKAIEEKKYLAGA DPSTVEMCYPPIIQSGGNYNLKFSVVSDKNHMHFGAITCAMGIRFKSYCSNLVRTLMVDPSQEVQENYNFLLQLQEELLK ELRHGVKICDVYNAVMDVVKKQKPELLNKITKNLGFGMGIEFREGSLVINSKNQYKLKKGMVFSINLGFSDLTNKEGKKP EEKTYALFIGDTVLVDEDGPATVLTSVKKKVKNVGIFLKNEDEEEEEEEKDEAEDLLGRGSRAALLTERTRNEMTAEEKR RAHQKELAAQLNEEAKRRLTEQKGEQQIQKARKSNVSYKNPSLMPKEPHIREMKIYIDKKYETVIMPVFGIATPFHIATI KNISMSVEGDYTYLRINFYCPGSALGRNEGNIFPNPEATFVKEITYRASNIKAPGEQTVPALNLQNAFRIIKEVQKRYKT REAEEKEKEGIVKQDSLVINLNRSNPKLKDLYIRPNIAQKRMQGSLEAHVNGFRFTSVRGDKVDILYNNIKHALFQPCDG EMIIVLHFHLKNAIMFGKKRHTDVQFYTEVGEITTDLGKHQHMHDRDDLYAEQMEREMRHKLKTAFKNFIEKVEALTKEE LEFEVPFRDLGFNGAPYRSTCLLQPTSSALVNATEWPPFVVTLDEVELIHFERVQFHLKNFDMVIVYKDYSKKVTMINAI PVASLDPIKEWLNSCDLKYTEGVQSLNWTKIMKTIVDDPEGFFEQGGWSFLEPEGEGSDAEEGDSESEIEDETFNPSEDD YEEEEEDSDEDYSSEAEESDYSKESLGSEEESGKDWDELEEEARKADRESRYEEEEEQSRSMSRKRKASVHSSGRGSNRG SRHSSAPPKKKRK Source: (gene. exp.) Homo sapiens (human) / Production host: Spodoptera frugiperda (fall armyworm) / References: UniProt: Q9Y5B9*PLUS |
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#6: Protein | Mass: 73361.172 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Details: The poly-(UNK) was placed to show the electron density between 171-198. The full sequence for the experiment is: ...Details: The poly-(UNK) was placed to show the electron density between 171-198. The full sequence for the experiment is: MAETLEFNDVYQEVKGSMNDGRLRLSRQGIIFKNSKTGKVDNIQAGELTEGIWRRVALGHGLKLLTKNGHVYKYDGFRES EFEKLSDFFKTHYRLELMEKDLCVKGWNWGTVKFGGQLLSFDIGDQPVFEIPLSNVSQCTTGKNEVTLEFHQNDDAEVSL MEVRFYVPPTQEDGVDPVEAFAQNVLSKADVIQATGDAICIFRELQCLTPRGRYDIRIYPTFLHLHGKTFDYKIPYTTVL RLFLLPHKDQRQMFFVISLDPPIKQGQTRYHFLILLFSKDEDISLTLNMNEEEVEKRFEGRLTKNMSGSLYEMVSRVMKA LVNRKITVPGNFQGHSGAQCITCSYKASSGLLYPLERGFIYVHKPPVHIRFDEISFVNFARGTTTTRSFDFEIETKQGTQ YTFSSIEREEYGKLFDFVNAKKLNIKNRGLKEGMNPSYDEYADSDEDQHDAYLERMKEEGKIREENANDSSDDSGEETDE SFNPGEEEEDVAEEFDSNASASSSSNEGDSDRDEKKRKQLKKAKMAKDRKSRKKPVEVKKGKDPNAPKRPMSAYMLWLNA SREKIKSDHPGISITDLSKKAGEIWKGMSKEKKEEWDRKAEDARRDYEKAMKEYEGGRGESSKRDKSKKKKKVKVKMEKK STPSRGSSSKSSSRQLSESFKSKEFVSSDESSSGENKSKKKRRRSEDSEEEELASTPPSSEDSASGSDE Source: (gene. exp.) Homo sapiens (human) / Production host: Spodoptera frugiperda (fall armyworm) / References: UniProt: Q08945*PLUS |
-DNA chain , 2 types, 2 molecules IJ
#7: DNA chain | Mass: 24144.422 Da / Num. of mol.: 1 / Source method: obtained synthetically / Source: (synth.) Escherichia coli (E. coli) |
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#8: DNA chain | Mass: 24584.680 Da / Num. of mol.: 1 / Source method: obtained synthetically / Source: (synth.) Escherichia coli (E. coli) |
-Experimental details
-Experiment
Experiment | Method: ELECTRON MICROSCOPY |
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EM experiment | Aggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction |
-Sample preparation
Component | Name: FACT_subNucleosome_complex_class2 / Type: COMPLEX / Entity ID: all / Source: RECOMBINANT |
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Molecular weight | Value: 0.35 MDa / Experimental value: NO |
Source (natural) | Organism: Homo sapiens (human) |
Source (recombinant) | Organism: Escherichia coli 'BL21-Gold(DE3)pLysS AG' (bacteria) |
Buffer solution | pH: 7.5 |
Specimen | Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES |
Vitrification | Cryogen name: ETHANE |
-Electron microscopy imaging
Experimental equipment | Model: Titan Krios / Image courtesy: FEI Company |
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Microscopy | Model: FEI TITAN KRIOS |
Electron gun | Electron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM |
Electron lens | Mode: BRIGHT FIELDBright-field microscopy |
Image recording | Electron dose: 7.8 e/Å2 / Film or detector model: GATAN K2 SUMMIT (4k x 4k) |
-Processing
Software | Name: PHENIX / Version: 1.12_2829: / Classification: refinement | ||||||||||||||||||||||||
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CTF correction | Type: NONE | ||||||||||||||||||||||||
3D reconstruction | Resolution: 7.4 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 6990 / Symmetry type: POINT | ||||||||||||||||||||||||
Refine LS restraints |
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