EMDB-12796: Cryo-EM structure of human RNA Polymerase I in complex with RRN3

Basic information

• Entry: Database: EMDB / ID: EMD-12796
• Title: Cryo-EM structure of human RNA Polymerase I in complex with RRN3
• Map data: Human Pol I in complex with RRN3 Map E, obtained by RELION focused refinement, unsharpened

Sample

• Complex: RNA polymerase I in complex with RRN3
  • Complex: RNA polymerase I
    • Protein or peptide: x 13 types
  • Complex: RNA polymerase I-specific transcription initiation factor RRN3
    • Protein or peptide: x 1 types
• Ligand: x 1 types

• Keywords: RNA polymerase I / human / rRNA transcription / DNA-dependent RNA polymerase / pre-initiation / RRN3 / TRANSCRIPTION

Function / homology

Function:

RNA polymerase I transcription regulator complex / negative regulation of protein localization to nucleolus / nucleologenesis / cytoplasm organization / neural crest formation / RNA polymerase I core binding / RNA Polymerase III Chain Elongation / RNA Polymerase III Transcription Termination / DNA/RNA hybrid binding / RNA polymerase I general transcription initiation factor activity / RNA polymerase I general transcription initiation factor binding / regulation of transcription by RNA polymerase I / negative regulation of intrinsic apoptotic signaling pathway by p53 class mediator / RNA polymerase I core promoter sequence-specific DNA binding / RPAP3/R2TP/prefoldin-like complex / RNA Polymerase III Transcription Initiation From Type 1 Promoter / RNA Polymerase III Transcription Initiation From Type 2 Promoter / RNA Polymerase III Transcription Initiation From Type 3 Promoter / Cytosolic sensors of pathogen-associated DNA / RNA Polymerase III Abortive And Retractive Initiation / RNA polymerase I preinitiation complex assembly / nucleolus organization / nucleobase-containing compound metabolic process / Abortive elongation of HIV-1 transcript in the absence of Tat / FGFR2 alternative splicing / RNA Polymerase I Transcription Termination / MicroRNA (miRNA) biogenesis / Viral Messenger RNA Synthesis / Signaling by FGFR2 IIIa TM / RNA Pol II CTD phosphorylation and interaction with CE during HIV infection / RNA Pol II CTD phosphorylation and interaction with CE / Formation of the Early Elongation Complex / Formation of the HIV-1 Early Elongation Complex / mRNA Capping / HIV Transcription Initiation / RNA Polymerase II HIV Promoter Escape / Transcription of the HIV genome / RNA Polymerase II Promoter Escape / RNA Polymerase II Transcription Pre-Initiation And Promoter Opening / RNA Polymerase II Transcription Initiation / RNA Polymerase II Transcription Initiation And Promoter Clearance / regulation of DNA-templated transcription initiation / termination of RNA polymerase I transcription / nucleolar large rRNA transcription by RNA polymerase I / mRNA Splicing - Minor Pathway / PIWI-interacting RNA (piRNA) biogenesis / transcription initiation at RNA polymerase I promoter / rRNA transcription / intrinsic apoptotic signaling pathway by p53 class mediator / RNA Polymerase I Transcription Initiation / Pausing and recovery of Tat-mediated HIV elongation / Tat-mediated HIV elongation arrest and recovery / Processing of Capped Intron-Containing Pre-mRNA / RNA polymerase II transcribes snRNA genes / HIV elongation arrest and recovery / Pausing and recovery of HIV elongation / homeostasis of number of cells / Tat-mediated elongation of the HIV-1 transcript / Formation of HIV-1 elongation complex containing HIV-1 Tat / transcription by RNA polymerase III / transcription by RNA polymerase I / RNA polymerase I complex / transcription elongation by RNA polymerase I / Formation of HIV elongation complex in the absence of HIV Tat / RNA polymerase III complex / RNA polymerase II, core complex / tRNA transcription by RNA polymerase III / RNA Polymerase II Transcription Elongation / : / Formation of RNA Pol II elongation complex / RNA Polymerase II Pre-transcription Events / embryo implantation / DNA-directed RNA polymerase activity / Inhibition of DNA recombination at telomere / mRNA Splicing - Major Pathway / cellular response to leukemia inhibitory factor / cell surface receptor protein tyrosine kinase signaling pathway / TP53 Regulates Transcription of DNA Repair Genes / RNA Polymerase I Promoter Escape / DNA-templated transcription initiation / Transcriptional regulation by small RNAs / protein-DNA complex / NoRC negatively regulates rRNA expression / B-WICH complex positively regulates rRNA expression / Transcription-Coupled Nucleotide Excision Repair (TC-NER) / ribonucleoside binding / Formation of TC-NER Pre-Incision Complex / Activation of anterior HOX genes in hindbrain development during early embryogenesis / : / : / : / fibrillar center / : / : / : / DNA-directed RNA polymerase / Dual incision in TC-NER / Gap-filling DNA repair synthesis and ligation in TC-NER / ribosome biogenesis / single-stranded DNA binding

Domain/homology:

RNA polymerase I specific transcription initiation factor RRN3 / RNA polymerase I specific transcription initiation factor RRN3 / DNA-directed RNA polymerase I, subunit RPA34.5 / DNA-directed RNA polymerase I subunit RPA34.5 / RNA polymerase I associated factor, A49-like / A49-like RNA polymerase I associated factor / Rpa43, N-terminal ribonucleoprotein (RNP) domain / Pol I subunit A12, C-terminal zinc ribbon / RPA43, OB domain / RPA43 OB domain in RNA Pol I / DNA-directed RNA polymerase I subunit RPA2, domain 4 / DNA-directed RNA pol I, largest subunit / : / RNA polymerase I, Rpa2 specific domain / DNA-directed RNA polymerases I and III subunit AC19 / DNA-directed RNA polymerases I and III subunit AC40 / Zinc finger TFIIS-type signature. / RNA polymerase Rpb7-like , N-terminal / RNA polymerase Rpb7-like, N-terminal domain superfamily / RNA polymerase subunit Rpb7-like / SHS2 domain found in N terminus of Rpb7p/Rpc25p/MJ0397 / DNA-directed RNA polymerase M, 15kDa subunit, conserved site / RNA polymerases M / 15 Kd subunits signature. / DNA-directed RNA polymerase subunit/transcription factor S / : / RNA polymerase, Rpb8 / DNA-directed RNA polymerases I, II, and III subunit RPABC4 / RNA polymerase Rpb8 / RNA polymerase subunit 8 / RNA polymerase, Rpb5, N-terminal / RNA polymerase Rpb5, N-terminal domain superfamily / RNA polymerase Rpb5, N-terminal domain / DNA-directed RNA polymerase, subunit RPB6 / DNA-directed RNA polymerase subunit RPABC5/Rpb10 / RNA polymerases, subunit N, zinc binding site / RNA polymerase subunit RPB10 / RNA polymerases N / 8 kDa subunit / RNA polymerases N / 8 Kd subunits signature. / DNA directed RNA polymerase, 7 kDa subunit / RNA polymerase archaeal subunit P/eukaryotic subunit RPABC4 / RNA polymerase, subunit H/Rpb5, conserved site / RNA polymerases H / 23 Kd subunits signature. / RNA polymerase subunit CX / DNA-directed RNA polymerase, 30-40kDa subunit, conserved site / DNA-directed RNA polymerase subunit Rpo3/Rpb3/RPAC1 / RNA polymerases D / 30 to 40 Kd subunits signature. / DNA-directed RNA polymerase Rpb11, 13-16kDa subunit, conserved site / DNA-directed RNA polymerase subunit Rpo11 / RNA polymerases L / 13 to 16 Kd subunits signature. / Zinc finger, TFIIS-type / Transcription factor S-II (TFIIS) / Zinc finger TFIIS-type profile. / C2C2 Zinc finger / RNA polymerase subunit RPABC4/transcription elongation factor Spt4 / DNA-directed RNA polymerase, RBP11-like dimerisation domain / RNA polymerase Rpb3/Rpb11 dimerisation domain / RNA polymerase, subunit H/Rpb5 C-terminal / DNA-directed RNA polymerase subunit Rpo5/Rpb5 / RPB5-like RNA polymerase subunit superfamily / RNA polymerase Rpb5, C-terminal domain / Archaeal Rpo6/eukaryotic RPB6 RNA polymerase subunit / DNA-directed RNA polymerase, 14-18kDa subunit, conserved site / RNA polymerases K / 14 to 18 Kd subunits signature. / DNA-directed RNA polymerase, subunit beta-prime / RNA polymerase Rpb6 / RNA polymerase, subunit omega/Rpo6/RPB6 / RNA polymerase Rpb6 / RNA polymerase Rpb2, domain 2 superfamily / RNA polymerase Rpb1, domain 3 superfamily / RPB6/omega subunit-like superfamily / DNA-directed RNA polymerase, insert domain / DNA-directed RNA polymerase, RpoA/D/Rpb3-type / RNA polymerase Rpb3/RpoA insert domain / RNA polymerase Rpb3/Rpb11 dimerisation domain / RNA polymerases D / RNA polymerase Rpb1, clamp domain superfamily / RNA polymerase Rpb1, domain 3 / RNA polymerase Rpb1, domain 3 / RNA polymerase Rpb1, domain 1 / RNA polymerase Rpb1, domain 1 / RNA polymerase, beta subunit, protrusion / RNA polymerase beta subunit / RNA polymerase, alpha subunit / RNA polymerase Rpb1, domain 5 / RNA polymerase Rpb1, domain 4 / RNA polymerase Rpb1, domain 2 / RNA polymerase Rpb1, domain 5 / RNA polymerase Rpb1, domain 4 / DNA-directed RNA polymerase, insert domain superfamily / RNA polymerase, N-terminal / RNA polymerase Rpb1, funnel domain superfamily / RNA polymerase I subunit A N-terminus / RNA polymerase Rpb2, domain 2 / RNA polymerase Rpb2, domain 2 / RNA polymerase, RBP11-like subunit / RNA polymerase, beta subunit, conserved site / RNA polymerase Rpb2, domain 7 / RNA polymerase Rpb2, domain 3 / RNA polymerase Rpb2, OB-fold / RNA polymerase Rpb2, domain 7

Component:

DNA-directed RNA polymerases I and III subunit RPAC1 / DNA-directed RNA polymerase I subunit RPA34 / DNA-directed RNA polymerase I subunit RPA1 / DNA-directed RNA polymerases I and III subunit RPAC2 / DNA-directed RNA polymerases I, II, and III subunit RPABC1 / DNA-directed RNA polymerases I, II, and III subunit RPABC3 / DNA-directed RNA polymerases I, II, and III subunit RPABC4 / DNA-directed RNA polymerases I, II, and III subunit RPABC2 / DNA-directed RNA polymerases I, II, and III subunit RPABC5 / DNA-directed RNA polymerase I subunit RPA43 / DNA-directed RNA polymerase I subunit RPA49 / DNA-directed RNA polymerase I subunit RPA2 / RNA polymerase I-specific transcription initiation factor RRN3 / DNA-directed RNA polymerase I subunit RPA12

• Biological species: Homo sapiens (human)
• Method: single particle reconstruction / cryo EM / Resolution: 3.1 Å
• Authors: Misiaszek AD / Girbig M
• Citation: Journal: Nat Struct Mol Biol / Year: 2021; Title: Cryo-EM structures of human RNA polymerase I.; Authors: Agata D Misiaszek / Mathias Girbig / Helga Grötsch / Florence Baudin / Brice Murciano / Aleix Lafita / Christoph W Müller / ; Abstract: RNA polymerase I (Pol I) specifically synthesizes ribosomal RNA. Pol I upregulation is linked to cancer, while mutations in the Pol I machinery lead to developmental disorders. Here we report the cryo-EM structure of elongating human Pol I at 2.7 Å resolution. In the exit tunnel, we observe a double-stranded RNA helix that may support Pol I processivity. Our structure confirms that human Pol I consists of 13 subunits with only one subunit forming the Pol I stalk. Additionally, the structure of human Pol I in complex with the initiation factor RRN3 at 3.1 Å resolution reveals stalk flipping upon RRN3 binding. We also observe an inactivated state of human Pol I bound to an open DNA scaffold at 3.3 Å resolution. Lastly, the high-resolution structure of human Pol I allows mapping of disease-related mutations that can aid understanding of disease etiology.

History

Deposition	Apr 21, 2021	-
Header (metadata) release	Dec 8, 2021	-
Map release	Dec 8, 2021	-
Update	Jul 10, 2024	-
Current status	Jul 10, 2024	Processing site: PDBe / Status: Released

Map

• File: Download / File: emd_12796.map.gz / Format: CCP4 / Size: 91.1 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
• Annotation: Human Pol I in complex with RRN3 Map E, obtained by RELION focused refinement, unsharpened
• Voxel size: X=Y=Z: 0.822 Å

Density

Contour Level	By AUTHOR: 0.00386 / Movie #1: 0.00386
Minimum - Maximum	-0.008483952 - 0.026302744
Average (Standard dev.)	0.00020343134 (±0.0014985278)

• Symmetry: Space group: 1

Details

EMDB XML:

Map geometry	Axis order X Y Z Origin 0 0 0 Dimensions 288 288 288 Spacing 288 288 288
Cell	A=B=C: 236.73601 Å α=β=γ: 90.0 °

CCP4 map header:

mode	Image stored as Reals
Å/pix. X/Y/Z	0.822	0.822	0.822
M x/y/z	288	288	288
origin x/y/z	0.000	0.000	0.000
length x/y/z	236.736	236.736	236.736
α/β/γ	90.000	90.000	90.000
MAP C/R/S	1	2	3
start NC/NR/NS	0	0	0
NC/NR/NS	288	288	288
D min/max/mean	-0.008	0.026	0.000

Supplemental data

Mask #1

• File: emd_12796_msk_1.map

Mask #2

• File: emd_12796_msk_2.map

Additional map: Human Pol I in complex with RRN3 Map...

• File: emd_12796_additional_1.map
• Annotation: Human Pol I in complex with RRN3 Map E, obtained by RELION focused refinement, sharpened with LocScale

Additional map: Human Pol I in complex with RRN3 Map...

• File: emd_12796_additional_2.map
• Annotation: Human Pol I in complex with RRN3 Map D, obtained by RELION masked classification, unsharpened

Additional map: Human Pol I in complex with RRN3 Map...

• File: emd_12796_additional_3.map
• Annotation: Human Pol I in complex with RRN3 Map D, obtained by RELION masked classification, sharpened with LocalDeblur

Additional map: Human Pol I in complex with RRN3 Map...

• File: emd_12796_additional_4.map
• Annotation: Human Pol I in complex with RRN3 Map E, obtained by RELION focused refinement, sharpened with LocalDeblur

Additional map: Human Pol I in complex with RRN3 Map...

• File: emd_12796_additional_5.map
• Annotation: Human Pol I in complex with RRN3 Map D, obtained by RELION masked classification, sharpened with LocScale

Half map: Human Pol I in complex with RRN3 Map E, half-map 2

• File: emd_12796_half_map_1.map
• Annotation: Human Pol I in complex with RRN3 Map E, half-map 2

Half map: Human Pol I in complex with RRN3 Map...

• File: emd_12796_half_map_2.map
• Annotation: Human Pol I in complex with RRN3 Map E, obtained by RELION focused refinement, unsharpened

Sample components

Entire : RNA polymerase I in complex with RRN3

• Entire: Name: RNA polymerase I in complex with RRN3

Components

• Complex: RNA polymerase I in complex with RRN3
  • Complex: RNA polymerase I
    • Protein or peptide: DNA-directed RNA polymerase I subunit RPA1
    • Protein or peptide: DNA-directed RNA polymerase I subunit RPA2
    • Protein or peptide: DNA-directed RNA polymerases I and III subunit RPAC1
    • Protein or peptide: DNA-directed RNA polymerases I, II, and III subunit RPABC1
    • Protein or peptide: DNA-directed RNA polymerases I, II, and III subunit RPABC2
    • Protein or peptide: DNA-directed RNA polymerase I subunit RPA43
    • Protein or peptide: DNA-directed RNA polymerases I, II, and III subunit RPABC3
    • Protein or peptide: DNA-directed RNA polymerase I subunit RPA12
    • Protein or peptide: DNA-directed RNA polymerases I, II, and III subunit RPABC5
    • Protein or peptide: DNA-directed RNA polymerases I and III subunit RPAC2
    • Protein or peptide: DNA-directed RNA polymerases I, II, and III subunit RPABC4
    • Protein or peptide: DNA-directed RNA polymerase I subunit RPA49
    • Protein or peptide: DNA-directed RNA polymerase I subunit RPA34
  • Complex: RNA polymerase I-specific transcription initiation factor RRN3
    • Protein or peptide: RNA polymerase I-specific transcription initiation factor RRN3
• Ligand: ZINC ION

Supramolecule #1: RNA polymerase I in complex with RRN3

• Supramolecule: Name: RNA polymerase I in complex with RRN3 / type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1-#14
• Molecular weight: Theoretical: 670 KDa

Supramolecule #2: RNA polymerase I

• Supramolecule: Name: RNA polymerase I / type: complex / ID: 2 / Parent: 1 / Macromolecule list: #1-#13
• Source (natural): Organism: Homo sapiens (human)

Supramolecule #3: RNA polymerase I-specific transcription initiation factor RRN3

• Supramolecule: Name: RNA polymerase I-specific transcription initiation factor RRN3; type: complex / ID: 3 / Parent: 1 / Macromolecule list: #14
• Source (natural): Organism: Homo sapiens (human)

Macromolecule #1: DNA-directed RNA polymerase I subunit RPA1

• Macromolecule: Name: DNA-directed RNA polymerase I subunit RPA1 / type: protein_or_peptide / ID: 1 / Number of copies: 1 / Enantiomer: LEVO / EC number: DNA-directed RNA polymerase
• Source (natural): Organism: Homo sapiens (human)
• Molecular weight: Theoretical: 195.069047 KDa

Sequence

String:

MLISKNMPWR RLQGISFGMY SAEELKKLSV KSITNPRYLD SLGNPSANGL YDLALGPADS KEVCSTCVQD FSNCSGHLGH IELPLTVYN PLLFDKLYLL LRGSCLNCHM LTCPRAVIHL LLCQLRVLEV GALQAVYELE RILNRFLEEN PDPSASEIRE E LEQYTTEI VQNNLLGSQG AHVKNVCESK SKLIALFWKA HMNAKRCPHC KTGRSVVRKE HNSKLTITFP AMVHRTAGQK DS EPLGIEE AQIGKRGYLT PTSAREHLSA LWKNEGFFLN YLFSGMDDDG MESRFNPSVF FLDFLVVPPS RYRPVSRLGD QMF TNGQTV NLQAVMKDVV LIRKLLALMA QEQKLPEEVA TPTTDEEKDS LIAIDRSFLS TLPGQSLIDK LYNIWIRLQS HVNI VFDSE MDKLMMDKYP GIRQILEKKE GLFRKHMMGK RVDYAARSVI CPDMYINTNE IGIPMVFATK LTYPQPVTPW NVQEL RQAV INGPNVHPGA SMVINEDGSR TALSAVDMTQ REAVAKQLLT PATGAPKPQG TKIVCRHVKN GDILLLNRQP TLHRPS IQA HRARILPEEK VLRLHYANCK AYNADFDGDE MNAHFPQSEL GRAEAYVLAC TDQQYLVPKD GQPLAGLIQD HMVSGAS MT TRGCFFTREH YMELVYRGLT DKVGRVKLLS PSILKPFPLW TGKQVVSTLL INIIPEDHIP LNLSGKAKIT GKAWVKET P RSVPGFNPDS MCESQVIIRE GELLCGVLDK AHYGSSAYGL VHCCYEIYGG ETSGKVLTCL ARLFTAYLQL YRGFTLGVE DILVKPKADV KRQRIIEEST HCGPQAVRAA LNLPEAASYD EVRGKWQDAH LGKDQRDFNM IDLKFKEEVN HYSNEINKAC MPFGLHRQF PENSLQMMVQ SGAKGSTVNT MQISCLLGQI ELEGRRPPLM ASGKSLPCFE PYEFTPRAGG FVTGRFLTGI K PPEFFFHC MAGREGLVDT AVKTSRSGYL QRCIIKHLEG LVVQYDLTVR DSDGSVVQFL YGEDGLDIPK TQFLQPKQFP FL ASNYEVI MKSQHLHEVL SRADPKKALH HFRAIKKWQS KHPNTLLRRG AFLSYSQKIQ EAVKALKLES ENRNGRSPGT QEM LRMWYE LDEESRRKYQ KKAAACPDPS LSVWRPDIYF ASVSETFETK VDDYSQEWAA QTEKSYEKSE LSLDRLRTLL QLKW QRSLC EPGEAVGLLA AQSIGEPSTQ MTLNTFHFAG RGEMNVTLGI PRLREILMVA SANIKTPMMS VPVLNTKKAL KRVKS LKKQ LTRVCLGEVL QKIDVQESFC MEEKQNKFQV YQLRFQFLPH AYYQQEKCLR PEDILRFMET RFFKLLMESI KKKNNK ASA FRNVNTRRAT QRDLDNAGEL GRSRGEQEGD EEEEGHIVDA EAEEGDADAS DAKRKEKQEE EVDYESEEEE EREGEEN DD EDMQEERNPH REGARKTQEQ DEEVGLGTEE DPSLPALLTQ PRKPTHSQEP QGPEAMERRV QAVREIHPFI DDYQYDTE E SLWCQVTVKL PLMKINFDMS SLVVSLAHGA VIYATKGITR CLLNETTNNK NEKELVLNTE GINLPELFKY AEVLDLRRL YSNDIHAIAN TYGIEAALRV IEKEIKDVFA VYGIAVDPRH LSLVADYMCF EGVYKPLNRF GIRSNSSPLQ QMTFETSFQF LKQATMLGS HDELRSPSAC LVVGKVVRGG TGLFELKQPL R

UniProtKB: DNA-directed RNA polymerase I subunit RPA1

Macromolecule #2: DNA-directed RNA polymerase I subunit RPA2

• Macromolecule: Name: DNA-directed RNA polymerase I subunit RPA2 / type: protein_or_peptide / ID: 2 / Number of copies: 1 / Enantiomer: LEVO / EC number: DNA-directed RNA polymerase
• Source (natural): Organism: Homo sapiens (human)
• Molecular weight: Theoretical: 128.379219 KDa

Sequence

String:

MDPGSRWRNL PSGPSLKHLT DPSYGIPREQ QKAALQELTR AHVESFNYAV HEGLGLAVQA IPPFEFAFKD ERISFTILDA VISPPTVPK GTICKEANVY PAECRGRRST YRGKLTADIN WAVNGISKGI IKQFLGYVPI MVKSKLCNLR NLPPQALIEH H EEAEEMGG YFIINGIEKV IRMLIMPRRN FPIAMIRPKW KTRGPGYTQY GVSMHCVREE HSAVNMNLHY LENGTVMLNF IY RKELFFL PLGFALKALV SFSDYQIFQE LIKGKEDDSF LRNSVSQMLR IVMEEGCSTQ KQVLNYLGEC FRVKLNVPDW YPN EQAAEF LFNQCICIHL KSNTEKFYML CLMTRKLFAL AKGECMEDNP DSLVNQEVLT PGQLFLMFLK EKLEGWLVSI KIAF DKKAQ KTSVSMNTDN LMRIFTMGID LTKPFEYLFA TGNLRSKTGL GLLQDSGLCV VADKLNFIRY LSHFRCVHRG ADFAK MRTT TVRRLLPESW GFLCPVHTPD GEPCGLMNHL TAVCEVVTQF VYTASIPALL CNLGVTPIDG APHRSYSECY PVLLDG VMV GWVDKDLAPG IADSLRHFKV LREKRIPPWM EVVLIPMTGK PSLYPGLFLF TTPCRLVRPV QNLALGKEEL IGTMEQI FM NVAIFEDEVF AGVTTHQELF PHSLLSVIAN FIPFSDHNQS PRNMYQCQMG KQTMGFPLLT YQDRSDNKLY RLQTPQSP L VRPSMYDYYD MDNYPIGTNA IVAVISYTGY DMEDAMIVNK ASWERGFAHG SVYKSEFIDL SEKIKQGDSS LVFGIKPGD PRVLQKLDDD GLPFIGAKLQ YGDPYYSYLN LNTGESFVMY YKSKENCVVD NIKVCSNDTG SGKFKCVCIT MRVPRNPTIG DKFASRHGQ KGILSRLWPA EDMPFTESGM VPDILFNPHG FPSRMTIGML IESMAGKSAA LHGLCHDATP FIFSEENSAL E YFGEMLKA AGYNFYGTER LYSGISGLEL EADIFIGVVY YQRLRHMVSD KFQVRTTGAR DRVTNQPIGG RNVQGGIRFG EM ERDALLA HGTSFLLHDR LFNCSDRSVA HVCVKCGSLL SPLLEKPPPS WSAMRNRKYN CTLCSRSDTI DTVSVPYVFR YFV AELAAM NIKVKLDVV

UniProtKB: DNA-directed RNA polymerase I subunit RPA2

Macromolecule #3: DNA-directed RNA polymerases I and III subunit RPAC1

• Macromolecule: Name: DNA-directed RNA polymerases I and III subunit RPAC1 / type: protein_or_peptide / ID: 3 / Number of copies: 1 / Enantiomer: LEVO
• Source (natural): Organism: Homo sapiens (human)
• Molecular weight: Theoretical: 39.301672 KDa

Sequence

String:

MAASQAVEEM RSRVVLGEFG VRNVHTTDFP GNYSGYDDAW DQDRFEKNFR VDVVHMDENS LEFDMVGIDA AIANAFRRIL LAEVPTMAV EKVLVYNNTS IVQDEILAHR LGLIPIHADP RLFEYRNQGD EEGTEIDTLQ FRLQVRCTRN PHAAKDSSDP N ELYVNHKV YTRHMTWIPL GNQADLFPEG TIRPVHDDIL IAQLRPGQEI DLLMHCVKGI GKDHAKFSPV ATASYRLLPD IT LLEPVEG EAAEELSRCF SPGVIEVQEV QGKKVARVAN PRLDTFSREI FRNEKLKKVV RLARVRDHYI FSVESTGVLP PDV LVSEAI KVLMGKCRRF LDELDAVQMD

UniProtKB: DNA-directed RNA polymerases I and III subunit RPAC1

Macromolecule #4: DNA-directed RNA polymerases I, II, and III subunit RPABC1

• Macromolecule: Name: DNA-directed RNA polymerases I, II, and III subunit RPABC1; type: protein_or_peptide / ID: 4 / Number of copies: 1 / Enantiomer: LEVO
• Source (natural): Organism: Homo sapiens (human)
• Molecular weight: Theoretical: 24.644318 KDa

Sequence

String:

MDDEEETYRL WKIRKTIMQL CHDRGYLVTQ DELDQTLEEF KAQFGDKPSE GRPRRTDLTV LVAHNDDPTD QMFVFFPEEP KVGIKTIKV YCQRMQEENI TRALIVVQQG MTPSAKQSLV DMAPKYILEQ FLQQELLINI TEHELVPEHV VMTKEEVTEL L ARYKLREN QLPRIQAGDP VARYFGIKRG QVVKIIRPSE TAGRYITYRL VQ

UniProtKB: DNA-directed RNA polymerases I, II, and III subunit RPABC1

Macromolecule #5: DNA-directed RNA polymerases I, II, and III subunit RPABC2

• Macromolecule: Name: DNA-directed RNA polymerases I, II, and III subunit RPABC2; type: protein_or_peptide / ID: 5 / Number of copies: 1 / Enantiomer: LEVO
• Source (natural): Organism: Homo sapiens (human)
• Molecular weight: Theoretical: 14.491026 KDa

Sequence

String:

MSDNEDNFDG DDFDDVEEDE GLDDLENAEE EGQENVEILP SGERPQANQK RITTPYMTKY ERARVLGTRA LQIAMCAPVM VELEGETDP LLIAMKELKA RKIPIIIRRY LPDGSYEDWG VDELIITD

UniProtKB: DNA-directed RNA polymerases I, II, and III subunit RPABC2

Macromolecule #6: DNA-directed RNA polymerase I subunit RPA43

• Macromolecule: Name: DNA-directed RNA polymerase I subunit RPA43 / type: protein_or_peptide / ID: 6 / Number of copies: 1 / Enantiomer: LEVO
• Source (natural): Organism: Homo sapiens (human)
• Molecular weight: Theoretical: 37.490379 KDa

Sequence

String:

MAAGCSEAPR PAAASDGSLV GQAGVLPCLE LPTYAAACAL VNSRYSCLVA GPHQRHIALS PRYLNRKRTG IREQLDAELL RYSESLLGV PIAYDNIKVV GELGDIYDDQ GHIHLNIEAD FVIFCPEPGQ KLMGIVNKVS SSHIGCLVHG CFNASIPKPE Q LSAEQWQT MEINMGDELE FEVFRLDSDA AGVFCIRGKL NITSLQFKRS EVSEEVTENG TEEAAKKPKK KKKKKDPETY EV DSGTTKL ADDADDTPME ESALQNTNNA NGIWEEEPKK KKKKKKHQEV QDQDPVFQGS DSSGYQSDHK KKKKKRKHSE EAE FTPPLK CSPKRKGKSN FL

UniProtKB: DNA-directed RNA polymerase I subunit RPA43

Macromolecule #7: DNA-directed RNA polymerases I, II, and III subunit RPABC3

• Macromolecule: Name: DNA-directed RNA polymerases I, II, and III subunit RPABC3; type: protein_or_peptide / ID: 7 / Number of copies: 1 / Enantiomer: LEVO
• Source (natural): Organism: Homo sapiens (human)
• Molecular weight: Theoretical: 17.162273 KDa

Sequence

String:

MAGILFEDIF DVKDIDPEGK KFDRVSRLHC ESESFKMDLI LDVNIQIYPV DLGDKFRLVI ASTLYEDGTL DDGEYNPTDD RPSRADQFE YVMYGKVYRI EGDETSTEAA TRLSAYVSYG GLLMRLQGDA NNLHGFEVDS RVYLLMKKLA F

UniProtKB: DNA-directed RNA polymerases I, II, and III subunit RPABC3

Macromolecule #8: DNA-directed RNA polymerase I subunit RPA12

• Macromolecule: Name: DNA-directed RNA polymerase I subunit RPA12 / type: protein_or_peptide / ID: 8 / Number of copies: 1 / Enantiomer: LEVO
• Source (natural): Organism: Homo sapiens (human)
• Molecular weight: Theoretical: 13.917695 KDa

Sequence

String:

MSVMDLANTC SSFQSDLDFC SDCGSVLPLP GAQDTVTCIR CGFNINVRDF EGKVVKTSVV FHQLGTAMPM SVEEGPECQG PVVDRRCPR CGHEGMAYHT RQMRSADEGQ TVFYTCTNCK FQEKEDS

UniProtKB: DNA-directed RNA polymerase I subunit RPA12

Macromolecule #9: DNA-directed RNA polymerases I, II, and III subunit RPABC5

• Macromolecule: Name: DNA-directed RNA polymerases I, II, and III subunit RPABC5; type: protein_or_peptide / ID: 9 / Number of copies: 1 / Enantiomer: LEVO
• Source (natural): Organism: Homo sapiens (human)
• Molecular weight: Theoretical: 7.655123 KDa

Sequence

String:

MIIPVRCFTC GKIVGNKWEA YLGLLQAEYT EGDALDALGL KRYCCRRMLL AHVDLIEKLL NYAPLEK

UniProtKB: DNA-directed RNA polymerases I, II, and III subunit RPABC5

Macromolecule #10: DNA-directed RNA polymerases I and III subunit RPAC2

• Macromolecule: Name: DNA-directed RNA polymerases I and III subunit RPAC2 / type: protein_or_peptide / ID: 10 / Number of copies: 1 / Enantiomer: LEVO
• Source (natural): Organism: Homo sapiens (human)
• Molecular weight: Theoretical: 15.259222 KDa

Sequence

String:

MEEDQELERK ISGLKTSMAE GERKTALEMV QAAGTDRHCV TFVLHEEDHT LGNSLRYMIM KNPEVEFCGY TTTHPSESKI NLRIQTRGT LPAVEPFQRG LNELMNVCQH VLDKFEASIK DYKDQKASRN ESTF

UniProtKB: DNA-directed RNA polymerases I and III subunit RPAC2

Macromolecule #11: DNA-directed RNA polymerases I, II, and III subunit RPABC4

• Macromolecule: Name: DNA-directed RNA polymerases I, II, and III subunit RPABC4; type: protein_or_peptide / ID: 11 / Number of copies: 1 / Enantiomer: LEVO
• Source (natural): Organism: Homo sapiens (human)
• Molecular weight: Theoretical: 7.018244 KDa

Sequence

String:

MDTQKDVQPP KQQPMIYICG ECHTENEIKS RDPIRCRECG YRIMYKKRTK RLVVFDAR

UniProtKB: DNA-directed RNA polymerases I, II, and III subunit RPABC4

Macromolecule #12: DNA-directed RNA polymerase I subunit RPA49

• Macromolecule: Name: DNA-directed RNA polymerase I subunit RPA49 / type: protein_or_peptide / ID: 12 / Number of copies: 1 / Enantiomer: LEVO
• Source (natural): Organism: Homo sapiens (human)
• Molecular weight: Theoretical: 47.330234 KDa

Sequence

String:

MAAEVLPSAR WQYCGAPDGS QRAVLVQFSN GKLQSPGNMR FTLYENKDST NPRKRNQRIL AAETDRLSYV GNNFGTGALK CNTLCRHFV GILNKTSGQM EVYDAELFNM QPLFSDVSVE SELALESQTK TYREKMDSCI EAFGTTKQKR ALNTRRMNRV G NESLNRAV AKAAETIIDT KGVTALVSDA IHNDLQDDSL YLPPCYDDAA KPEDVYKFED LLSPAEYEAL QSPSEAFRNV TS EEILKMI EENSHCTFVI EALKSLPSDV ESRDRQARCI WFLDTLIKFR AHRVVKRKSA LGPGVPHIIN TKLLKHFTCL TYN NGRLRN LISDSMKAKI TAYVIILALH IHDFQIDLTV LQRDLKLSEK RMMEIAKAMR LKISKRRVSV AAGSEEDHKL GTLS LPLPP AQTSDRLAKR RKIT

UniProtKB: DNA-directed RNA polymerase I subunit RPA49

Macromolecule #13: DNA-directed RNA polymerase I subunit RPA34

• Macromolecule: Name: DNA-directed RNA polymerase I subunit RPA34 / type: protein_or_peptide / ID: 13 / Number of copies: 1 / Enantiomer: LEVO
• Source (natural): Organism: Homo sapiens (human)
• Molecular weight: Theoretical: 55.065523 KDa

Sequence

String:

MEEPQAGDAA RFSCPPNFTA KPPASESPRF SLEALTGPDT ELWLIQAPAD FAPECFNGRH VPLSGSQIVK GKLAGKRHRY RVLSSCPQA GEATLLAPST EAGGGLTCAS APQGTLRILE GPQQSLSGSP LQPIPASPPP QIPPGLRPRF CAFGGNPPVT G PRSALAPN LLTSGKKKKE MQVTEAPVTQ EAVNGHGALE VDMALGSPEM DVRKKKKKKN QQLKEPEAAG PVGTEPTVET LE PLGVLFP STTKKRKKPK GKETFEPEDK TVKQEQINTE PLEDTVLSPT KKRKRQKGTE GMEPEEGVTV ESQPQVKVEP LEE AIPLPP TKKRKKEKGQ MAMMEPGTEA MEPVEPEMKP LESPGGTMAP QQPEGAKPQA QAALAAPKKK TKKEKQQDAT VEPE TEVVG PELPDDLEPQ AAPTSTKKKK KKKERGHTVT EPIQPLEPEL PGEGQPEARA TPGSTKKRKK QSQESRMPET VPQEE MPGP PLNSESGEEA PTGRDKKRKQ QQQQPV

UniProtKB: DNA-directed RNA polymerase I subunit RPA34

Macromolecule #14: RNA polymerase I-specific transcription initiation factor RRN3

• Macromolecule: Name: RNA polymerase I-specific transcription initiation factor RRN3; type: protein_or_peptide / ID: 14 / Number of copies: 1 / Enantiomer: LEVO
• Source (natural): Organism: Homo sapiens (human)
• Molecular weight: Theoretical: 74.188031 KDa
• Recombinant expression: Organism: Escherichia coli (E. coli)

Sequence

String:

MAAPLLHTRL PGDAAASSSA VKKLGASRTG ISNMRALEND FFNSPPRKTV RFGGTVTEVL LKYKKGETND FELLKNQLLD PDIKDDQII NWLLEFRSSI MYLTKDFEQL ISIILRLPWL NRSQTVVEEY LAFLGNLVSA QTVFLRPCLS MIASHFVPPR V IIKEGDVD VSDSDDEDDN LPANFDTCHR ALQIIARYVP STPWFLMPIL VEKFPFVRKS ERTLECYVHN LLRISVYFPT LR HEILELI IEKLLKLDVN ASRQGIEDAE ETATQTCGGT DSTEGLFNMD EDEETEHETK AGPERLDQMV HPVAERLDIL MSL VLSYMK DVCYVDGKVD NGKTKDLYRD LINIFDKLLL PTHASCHVQF FMFYLCSFKL GFAEAFLEHL WKKLQDPSNP AIIR QAAGN YIGSFLARAK FIPLITVKSC LDLLVNWLHI YLNNQDSGTK AFCDVALHGP FYSACQAVFY TFVFRHKQLL SGNLK EGLQ YLQSLNFERI VMSQLNPLKI CLPSVVNFFA AITNKYQLVF CYTIIERNNR QMLPVIRSTA GGDSVQICTN PLDTFF PFD PCVLKRSKKF IDPIYQVWED MSAEELQEFK KPMKKDIVED EDDDFLKGEV PQNDTVIGIT PSSFDTHFRS PSSSVGS PP VLYMQPSPL

UniProtKB: RNA polymerase I-specific transcription initiation factor RRN3

Macromolecule #15: ZINC ION

• Macromolecule: Name: ZINC ION / type: ligand / ID: 15 / Number of copies: 7 / Formula: ZN
• Molecular weight: Theoretical: 65.409 Da

Experimental details

Structure determination

• Method: cryo EM
• Processing: single particle reconstruction
• Aggregation state: particle

Sample preparation

• Concentration: 0.85 mg/mL

Buffer

pH: 7.5
Component:

Concentration	Formula	Name
15.0 mM	C8H18N2O4S	HEPES
80.0 mM	(NH4)2SO4	ammonium sulfate
5.0 mM	MgCl2	magnesium chloride
10.0 mM	C4H10O2S2	dithiothreitol

• Grid: Model: Quantifoil R2/1 / Material: COPPER / Mesh: 200 / Pretreatment - Type: PLASMA CLEANING / Pretreatment - Time: 30 sec. / Pretreatment - Atmosphere: OTHER; Details: NanoClean plasma cleaner (Fischione Instruments, Model 1070)
• Vitrification: Cryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 288 K / Instrument: FEI VITROBOT MARK IV / Details: blot force 3, blot time 0 s, wait time 0 s.
• Details: Human RNA polymerase I mixed in 1:1 molar ratio with RRN3 and in 1:1.5 molar ratio with DNA template

Electron microscopy

• Microscope: FEI TITAN KRIOS
• Image recording: Film or detector model: GATAN K3 (6k x 4k) / Number grids imaged: 1 / Number real images: 14224 / Average electron dose: 41.2 e/Ų
• Electron beam: Acceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
• Electron optics: C2 aperture diameter: 100.0 µm / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: 2.25 µm / Nominal defocus min: 0.75 µm / Nominal magnification: 105000
• Sample stage: Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
• Experimental equipment: Model: Titan Krios / Image courtesy: FEI Company

Image processing

• Particle selection: Number selected: 2628144
• Startup model: Type of model: NONE
• Final reconstruction: Number classes used: 1 / Resolution.type: BY AUTHOR / Resolution: 3.1 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION (ver. 3.1) / Number images used: 260363
• Initial angle assignment: Type: MAXIMUM LIKELIHOOD / Software: (Name: cryoSPARC (ver. 3.1), RELION (ver. 3.1))
• Final angle assignment: Type: MAXIMUM LIKELIHOOD / Software - Name: RELION (ver. 3.1)
• Final 3D classification: Number classes: 2 / Avg.num./class: 200000 / Software - Name: RELION (ver. 3.1); Details: Masked classification was used at the final stages of processing

Atomic model buiding 1

Initial model

PDB ID	Chain
7aei	chain_id: C, source_name: PDB, initial_model_type: experimental model
7aei	chain_id: E, source_name: PDB, initial_model_type: experimental model
7aei	chain_id: F, source_name: PDB, initial_model_type: experimental model
7aei	chain_id: H, source_name: PDB, initial_model_type: experimental model
7aei	chain_id: J, source_name: PDB, initial_model_type: experimental model
7aei	chain_id: K, source_name: PDB, initial_model_type: experimental model
7aei	chain_id: L, source_name: PDB, initial_model_type: experimental model

• Details: Initial rigid body fitting with UCSF Chimera followed by manual model fitting in Coot.

Output model

PDB-7oba:
Cryo-EM structure of human RNA Polymerase I in complex with RRN3