EMDB-12971: Structure of active transcription elongation complex Pol II-DSIF-...

Basic information

• Entry: Database: EMDB / ID: EMD-12971
• Title: Structure of active transcription elongation complex Pol II-DSIF-ELL2-EAF1, Map 9, Global map used to fit SPT4-SPT5-NGN
• Map data: Structure of active transcription elongation complex Pol II-DSIF-ELL2-EAF1, Map 9, Global map used to fit SPT4-SPT5-NGN

Sample

• Complex: Transcription elongation complex of RNA polymerase II with elongation factors DSIF, ELL2 and EAF1
  • Complex: DNA-directed RNA polymerase
    • Protein or peptide: x 12 types
  • Complex: Elongation factors
    • Protein or peptide: x 2 types
  • Complex: RNA
    • DNA: x 2 types
    • RNA: x 1 types
  • Complex: Elongation factors
    • Protein or peptide: x 2 types

Function / homology

Function:

super elongation complex / negative regulation of DNA-templated transcription, elongation / : / DSIF complex / snRNA transcription by RNA polymerase II / regulation of transcription elongation by RNA polymerase II / B-WICH complex positively regulates rRNA expression / RNA Polymerase I Transcription Initiation / RNA Polymerase I Promoter Escape / RNA Polymerase I Transcription Termination / 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 / Formation of RNA Pol II elongation complex / Formation of the Early Elongation Complex / Transcriptional regulation by small RNAs / RNA Polymerase II Pre-transcription Events / TP53 Regulates Transcription of DNA Repair Genes / FGFR2 alternative splicing / RNA polymerase II transcribes snRNA genes / mRNA Capping / mRNA Splicing - Minor Pathway / Processing of Capped Intron-Containing Pre-mRNA / RNA Polymerase II Promoter Escape / RNA Polymerase II Transcription Pre-Initiation And Promoter Opening / RNA Polymerase II Transcription Initiation / RNA Polymerase II Transcription Elongation / RNA Polymerase II Transcription Initiation And Promoter Clearance / RNA Pol II CTD phosphorylation and interaction with CE / Estrogen-dependent gene expression / Formation of TC-NER Pre-Incision Complex / Dual incision in TC-NER / Gap-filling DNA repair synthesis and ligation in TC-NER / mRNA Splicing - Major Pathway / transcription elongation factor activity / positive regulation of DNA-templated transcription, elongation / Abortive elongation of HIV-1 transcript in the absence of Tat / transcription elongation-coupled chromatin remodeling / RNA Pol II CTD phosphorylation and interaction with CE during HIV infection / RNA Pol II CTD phosphorylation and interaction with CE / intercellular bridge / Formation of the Early Elongation Complex / Formation of the HIV-1 Early Elongation Complex / mRNA Capping / organelle membrane / maintenance of transcriptional fidelity during transcription elongation by RNA polymerase II / RNA polymerase III activity / negative regulation of transcription elongation by RNA polymerase II / RNA polymerase II complex binding / Pausing and recovery of Tat-mediated HIV elongation / Tat-mediated HIV elongation arrest and recovery / positive regulation of macroautophagy / positive regulation of nuclear-transcribed mRNA poly(A) tail shortening / HIV elongation arrest and recovery / Pausing and recovery of HIV elongation / RNA polymerase II transcribes snRNA genes / RNA polymerase II activity / Tat-mediated elongation of the HIV-1 transcript / transcription elongation by RNA polymerase I / Formation of HIV-1 elongation complex containing HIV-1 Tat / transcription-coupled nucleotide-excision repair / tRNA transcription by RNA polymerase III / RNA polymerase I complex / RNA polymerase I activity / RNA polymerase III complex / cis-regulatory region sequence-specific DNA binding / positive regulation of translational initiation / Formation of HIV elongation complex in the absence of HIV Tat / RNA polymerase II, core complex / Cajal body / RNA Polymerase II Transcription Elongation / Formation of RNA Pol II elongation complex / RNA Polymerase II Pre-transcription Events / translation initiation factor binding / positive regulation of RNA splicing / transcription elongation factor complex / promoter-specific chromatin binding / TP53 Regulates Transcription of DNA Repair Genes / transcription initiation at RNA polymerase II promoter / positive regulation of transcription elongation by RNA polymerase II / transcription elongation by RNA polymerase II / P-body / ribonucleoside binding / fibrillar center / DNA-directed 5'-3' RNA polymerase activity / DNA-directed RNA polymerase / single-stranded DNA binding / chromosome / Hydrolases; Acting on ester bonds; Exoribonucleases producing 5'-phosphomonoesters / transcription by RNA polymerase II / nucleic acid binding / chromosome, telomeric region / single-stranded RNA binding / protein dimerization activity / nuclear body / hydrolase activity / nuclear speck / protein heterodimerization activity / RNA-directed RNA polymerase / intracellular membrane-bounded organelle

Domain/homology:

Transcription elongation factor Eaf, N-terminal / EAF family / RNA polymerase II transcription elongation factor / RNA polymerase II elongation factor ELL, N-terminal / RNA polymerase II elongation factor ELL / Occludin homology domain / ELL/occludin family / Occludin homology domain / Occludin/ELL (OCEL) domain profile. / E3 ubiquitin-protein ligase ELL-like / Spt5, KOW domain repeat 6 / Transcription initiation Spt4 / Spt4 superfamily / Spt4/RpoE2 zinc finger / Spt4/RpoE2 zinc finger / Spt4/RpoE2 zinc finger / Transcription elongation factor Spt5, eukaryote / Spt5 transcription elongation factor, N-terminal / Spt5, KOW domain repeat 2 / Spt5, KOW domain repeat 3 / Spt5, KOW domain repeat 5 / Spt5 transcription elongation factor, acidic N-terminal / NGN domain, eukaryotic / Spt5, KOW domain repeat 1 / Spt5, KOW domain repeat 4 / Spt5 C-terminal domain / Spt5 C-terminal nonapeptide repeat binding Spt4 / NGN domain / Transcription elongation factor SPT5 / Early transcription elongation factor of RNA pol II, NGN section / RNA-binding domain, S1 / NusG, N-terminal / In Spt5p, this domain may confer affinity for Spt4p. It possesses a RNP-like fold. / NusG, N-terminal domain superfamily / DNA-directed RNA polymerase II subunit Rpb4-like / RNA polymerase Rpb4/RPC9, core / DNA-directed RNA-polymerase II subunit / RNA polymerase II, heptapeptide repeat, eukaryotic / RNA polymerase Rpb1, domain 6 / RNA polymerase Rpb1, domain 6 / RNA polymerase Rpb1 C-terminal repeat / Eukaryotic RNA polymerase II heptapeptide repeat. / RNA polymerase Rpb1, domain 7 / RNA polymerase Rpb1, domain 7 superfamily / RNA polymerase Rpb1, domain 7 / Rpb4/RPC9 superfamily / Pol II subunit B9, C-terminal zinc ribbon / RNA polymerase RBP11 / RNA polymerase subunit Rpb4/RPC9 / RNA polymerase Rpb4 / Zinc finger TFIIS-type signature. / HRDC-like superfamily / 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 / RNA polymerase Rpb2, domain 5 / RNA polymerase Rpb2, domain 5 / RNA polymerase Rpb2, domain 4 / RNA polymerase Rpb2, domain 4 / DNA-directed RNA polymerase, M/15kDa subunit / RNA polymerases M/15 Kd subunit / RNA polymerase subunit 9 / 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 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, subunit RPB6 / 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 / DNA-directed RNA polymerase subunit Rpo5/Rpb5 / 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

Component:

RNA polymerase II subunit D / DNA-directed RNA polymerase II subunit RPB11-a / DNA-directed RNA polymerases I, II, and III subunit RPABC3 / DNA-directed RNA polymerases I, II, and III subunit RPABC5 / DNA-directed RNA polymerases I, II, and III subunit RPABC2 / DNA-directed RNA polymerase II subunit RPB3 / DNA-directed RNA polymerase subunit beta / DNA-directed RNA polymerase II subunit RPB7 / RNA polymerase II, I and III subunit K / DNA-directed RNA polymerase II subunit E / Transcription elongation factor SPT5 / RNA polymerase II elongation factor ELL2 / DNA-directed RNA polymerase II subunit RPB1 / DNA-directed RNA polymerase II subunit RPB9 / Transcription elongation factor SPT4 / ELL-associated factor 1

• Biological species: Sus scrofa (pig) / Homo sapiens (human) / HIV whole-genome vector AA1305#18 (others)
• Method: single particle reconstruction / cryo EM / Resolution: 3.5 Å
• Authors: Chen Y / Vos SM / Dienemann C / Ninov M / Urlaub H / Cramer P

Funding support

Germany, 4 items

Organization	Grant number	Country
German Research Foundation (DFG)	SFB860	Germany
German Research Foundation (DFG)	EXC 2067/1-390729940	Germany
European Research Council (ERC)	Advanced Grant CHROMATRANS 693023	Germany
German Research Foundation (DFG)	SFB1286	Germany

• Citation: Journal: Mol Cell / Year: 2021; Title: Allosteric transcription stimulation by RNA polymerase II super elongation complex.; Authors: Ying Chen / Seychelle M Vos / Christian Dienemann / Momchil Ninov / Henning Urlaub / Patrick Cramer / ; Abstract: The super elongation complex (SEC) contains the positive transcription elongation factor b (P-TEFb) and the subcomplex ELL2-EAF1, which stimulates RNA polymerase II (RNA Pol II) elongation. Here, we report the cryoelectron microscopy (cryo-EM) structure of ELL2-EAF1 bound to a RNA Pol II elongation complex at 2.8 Å resolution. The ELL2-EAF1 dimerization module directly binds the RNA Pol II lobe domain, explaining how SEC delivers P-TEFb to RNA Pol II. The same site on the lobe also binds the initiation factor TFIIF, consistent with SEC binding only after the transition from transcription initiation to elongation. Structure-guided functional analysis shows that the stimulation of RNA elongation requires the dimerization module and the ELL2 linker that tethers the module to the RNA Pol II protrusion. Our results show that SEC stimulates elongation allosterically and indicate that this stimulation involves stabilization of a closed conformation of the RNA Pol II active center cleft.

History

Deposition	May 18, 2021	-
Header (metadata) release	Jul 14, 2021	-
Map release	Jul 14, 2021	-
Update	Sep 1, 2021	-
Current status	Sep 1, 2021	Processing site: PDBe / Status: Released

Map

• File: Download / File: emd_12971.map.gz / Format: CCP4 / Size: 103 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
• Annotation: Structure of active transcription elongation complex Pol II-DSIF-ELL2-EAF1, Map 9, Global map used to fit SPT4-SPT5-NGN
• Voxel size: X=Y=Z: 1.05 Å

Density

Contour Level	By AUTHOR: 0.01 / Movie #1: 0.01
Minimum - Maximum	-0.020401707 - 0.055727407
Average (Standard dev.)	0.00024484753 (±0.002503212)

• Symmetry: Space group: 1

Details

EMDB XML:

Map geometry	Axis order X Y Z Origin 0 0 0 Dimensions 300 300 300 Spacing 300 300 300
Cell	A=B=C: 315.0 Å α=β=γ: 90.0 °

CCP4 map header:

mode	Image stored as Reals
Å/pix. X/Y/Z	1.05	1.05	1.05
M x/y/z	300	300	300
origin x/y/z	0.000	0.000	0.000
length x/y/z	315.000	315.000	315.000
α/β/γ	90.000	90.000	90.000
start NX/NY/NZ	0	0	0
NX/NY/NZ	640	640	640
MAP C/R/S	1	2	3
start NC/NR/NS	0	0	0
NC/NR/NS	300	300	300
D min/max/mean	-0.020	0.056	0.000

Supplemental data

Mask #1

• File: emd_12971_msk_1.map

Half map: half map 2 for map 9

• File: emd_12971_half_map_1.map
• Annotation: half map 2 for map 9

Half map: half map 1 for map 9

• File: emd_12971_half_map_2.map
• Annotation: half map 1 for map 9

Sample components

Entire : Transcription elongation complex of RNA polymerase II with elonga...

• Entire: Name: Transcription elongation complex of RNA polymerase II with elongation factors DSIF, ELL2 and EAF1

Components

• Complex: Transcription elongation complex of RNA polymerase II with elongation factors DSIF, ELL2 and EAF1
  • Complex: DNA-directed RNA polymerase
    • Protein or peptide: DNA-directed RNA polymerase II subunit RPB1
    • Protein or peptide: DNA-directed RNA polymerase II subunit beta
    • Protein or peptide: DNA-directed RNA polymerase II subunit RPB3
    • Protein or peptide: RNA polymerase II subunit D
    • Protein or peptide: RNA polymerase II subunit E
    • Protein or peptide: RNA polymerase II subunit F
    • Protein or peptide: RNA polymerase II subunit G
    • Protein or peptide: DNA-directed RNA polymerases I, II, and III subunit RPABC3
    • Protein or peptide: DNA-directed RNA polymerase II subunit RPB9
    • Protein or peptide: DNA-directed RNA polymerases I, II, and III subunit RPABC5
    • Protein or peptide: DNA-directed RNA polymerase II subunit RPB11-a
    • Protein or peptide: RNA polymerase II subunit K
  • Complex: Elongation factors
    • Protein or peptide: RNA polymerase II elongation factor ELL2
    • Protein or peptide: ELL-associated factor 1
  • Complex: RNA
    • DNA: Template DNA
    • DNA: Non-template DNA
    • RNA: RNA
  • Complex: Elongation factors
    • Protein or peptide: Transcription elongation factor SPT5
    • Protein or peptide: Transcription elongation factor SPT4

Supramolecule #1: Transcription elongation complex of RNA polymerase II with elonga...

• Supramolecule: Name: Transcription elongation complex of RNA polymerase II with elongation factors DSIF, ELL2 and EAF1; type: complex / ID: 1 / Parent: 0 / Macromolecule list: all

Supramolecule #2: DNA-directed RNA polymerase

• Supramolecule: Name: DNA-directed RNA polymerase / type: complex / ID: 2 / Parent: 1 / Macromolecule list: #1-#12
• Source (natural): Organism: Sus scrofa (pig)

Supramolecule #3: Elongation factors

• Supramolecule: Name: Elongation factors / type: complex / ID: 3 / Parent: 1 / Macromolecule list: #13-#14
• Source (natural): Organism: Homo sapiens (human)
• Recombinant expression: Organism: Trichoplusia ni (cabbage looper)

Supramolecule #4: RNA

• Supramolecule: Name: RNA / type: complex / ID: 4 / Parent: 1 / Macromolecule list: #17-#19
• Source (natural): Organism: HIV whole-genome vector AA1305#18 (others)
• Recombinant expression: Organism: synthetic construct (others)

Supramolecule #5: Elongation factors

• Supramolecule: Name: Elongation factors / type: complex / ID: 5 / Parent: 1 / Macromolecule list: #15-#16
• Source (natural): Organism: Homo sapiens (human)
• Recombinant expression: Organism: Escherichia coli BL21(DE3) (bacteria)

Macromolecule #1: DNA-directed RNA polymerase II subunit RPB1

• Macromolecule: Name: DNA-directed RNA polymerase II subunit RPB1 / type: protein_or_peptide / ID: 1 / Enantiomer: LEVO
• Source (natural): Organism: Sus scrofa (pig)

Sequence

String:

MHGGGPPSGD SACPLRTIKR VQFGVLSPDE LKRMSVTEGG IKYPETTEGG RPKLGGLMDP RQGVIERTGR CQTCAGNMT ECPGHFGHIE LAKPVFHVGF LVKTMKVLRC VCFFCSKLLV DSNNPKIKDI LAKSKGQPKK R LTHVYDLC KGKNICEGGE EMDNKFGVEQ PEGDEDLTKE KGHGGCGRYQ PRIRRSGLEL YAEWKHVNED SQ EKKILLS PERVHEIFKR ISDEECFVLG MEPRYARPEW MIVTVLPVPP LSVRPAVVMQ GSARNQDDLT HKL ADIVKI NNQLRRNEQN GAAAHVIAED VKLLQFHVAT MVDNELPGLP RAMQKSGRPL KSLKQRLKGK EGRV RGNLM GKRVDFSART VITPDPNLSI DQVGVPRSIA ANMTFAEIVT PFNIDRLQEL VRRGNSQYPG AKYII RDNG DRIDLRFHPK PSDLHLQTGY KVERHMCDGD IVIFNRQPTL HKMSMMGHRV RILPWSTFRL NLSVTT PYN ADFDGDEMNL HLPQSLETRA EIQELAMVPR MIVTPQSNRP VMGIVQDTLT AVRKFTKRDV FLERGEV MN LLMFLSTWDG KVPQPAILKP RPLWTGKQIF SLIIPGHINC IRTHSTHPDD EDSGPYKHIS PGDTKVVV E NGELIMGILC KKSLGTSAGS LVHISYLEMG HDITRLFYSN IQTVINNWLL IEGHTIGIGD SIADSKTYQ DIQNTIKKAK QDVIEVIEKA HNNELEPTPG NTLRQTFENQ VNRILNDARD KTGSSAQKSL SEYNNFKSMV VSGAKGSKI NISQVIAVVG QQNVEGKRIP FGFKHRTLPH FIKDDYGPES RGFVENSYLA GLTPTEFFFH A MGGREGLI DTAVKTAETG YIQRRLIKSM ESVMVKYDAT VRNSINQVVQ LRYGEDGLAG ESVEFQNLAT LK PSNKAFE KKFRFDYTNE RALRRTLQED LVKDVLSNAH IQNELEREFE RMREDREVLR VIFPTGDSKV VLP CNLLRM IWNAQKIFHI NPRLPSDLHP IKVVEGVKEL SKKLVIVNGD DPLSRQAQEN ATLLFNIHLR STLC SRRMA EEFRLSGEAF DWLLGEIESK FNQAIAHPGE MVGALAAQSL GEPATQMTLN TFHYAGVSAK NVTLG VPRL KELINISKKP KTPSLTVFLL GQSARDAERA KDILCRLEHT TLRKVTANTA IYYDPNPQST VVAEDQ EWV NVYYEMPDFD VARISPWLLR VELDRKHMTD RKLTMEQIAE KINAGFGDDL NCIFNDDNAE KLVLRIR IM NSDENKMQEE EEVVDKMDDD VFLRCIESNM LTDMTLQGIE QISKVYMHLP QTDNKKKIII TEDGEFKA L QEWILETDGV SLMRVLSEKD VDPVRTTSND IVEIFTVLGI EAVRKALERE LYHVISFDGS YVNYRHLAL LCDTMTCRGH LMAITRHGVN RQDTGPLMKC SFEETVDVLM EAAAHGESDP MKGVSENIML GQLAPAGTGC FDLLLDAEK CKYGMEIPTN IPGLGAAGPT GMFFGSAPSP MGGISPAMTP WNQGATPAYG AWSPSVGSGM T PGAAGFSP SAASDASGFS PGYSPAWSPT PGSPGSPGPS SPYIPSPGGA MSPSYSPTSP AYEPRSPGGY TP QSPSYSP TSPSYSPTSP SYSPTSPNYS PTSPSYSPTS PSYSPTSPSY SPTSPSYSPT SPSYSPTSPS YSP TSPSYS PTSPSYSPTS PSYSPTSPSY SPTSPSYSPT SPSYSPTSPS YSPTSPSYSP TSPSYSPTSP NYSP TSPNY TPTSPSYSPT SPSYSPTSPN YTPTSPNYSP TSPSYSPTSP SYSPTSPSYS PSSPRYTPQS PTYTP SSPS YSPSSPSYSP TSPKYTPTSP SYSPSSPEYT PTSPKYSPTS PKYSPTSPKY SPTSPTYSPT TPKYSP TSP TYSPTSPVYT PTSPKYSPTS PTYSPTSPKY SPTSPTYSPT SPKGSTYSPT SPGYSPTSPT YSLTSPA IS PDDSDEEN

Macromolecule #2: DNA-directed RNA polymerase II subunit beta

• Macromolecule: Name: DNA-directed RNA polymerase II subunit beta / type: protein_or_peptide / ID: 2 / Enantiomer: LEVO
• Source (natural): Organism: Sus scrofa (pig)

Sequence

String:

MCSTNLSQAL AYFRAGANLT AASLWAGFRG SRRIFWERRK RKSLCLALRP GGACWRWLLA VSCVSLRGL GAPGSCANMY DADEDMQYDE DDDEITPDLW QEACWIVISS YFDEKGLVRQ Q LDSFDEFI QMSVQRIVED APPIDLQAEA QHASGEVEEP PRYLLKFEQI YLSKPTHWER DG APSPMMP NEARLRNLTY SAPLYVDITK TVIKEGEEQL QTQHQKTFIG KIPIMLRSTY CLL NGLTDR DLCELNECPL DPGGYFIING SEKVLIAQEK MATNTVYVFA KKDSKYAYTG ECRS CLENS SRPTSTIWVS MLARGGQGAK KSAIGQRIVA TLPYIKQEVP IIIVFRALGF VSDRD ILEH IIYDFEDPEM MEMVKPSLDE AFVIQEQNVA LNFIGSRGAK PGVTKEKRIK YAKEVL QKE MLPHVGVSDF CETKKAYFLG YMVHRLLLAA LGRRELDDRD HYGNKRLDLA GPLLAFL FR GMFKNLLKEV RIYAQKFIDR GKDFNLELAI KTRIISDGLK YSLATGNWGD QKKAHQAR A GVSQVLNRLT FASTLSHLRR LNSPIGRDGK LAKPRQLHNT LWGMVCPAET PEGHAVGLV KNLALMAYIS VGSQPSPILE FLEEWSMENL EEISPAAIAD ATKIFVNGCW VGIHKDPEQL MNTLRKLRR QMDIIVSEVS MIRDIREREI RIYTDAGRIC RPLLIVEKQK LLLKKRHIDQ L KEREYNNY SWQDLVASGV VEYIDTLEEE TVMLAMTPDD LQEKEVAYCS TYTHCEIHPS MI LGVCASI IPFPDHNQSP RNTYQSAMGK QAMGVYITNF HVRMDTLAHV LYYPQKPLVT TRS MEYLRF RELPAGINSI VAIASYTGYN QEDSVIMNRS AVDRGFFRSV FYRSYKEQES KKGF DQEEV FEKPTRETCQ GMRHAIYDKL DDDGLIAPGV RVSGDDVIIG KTVTLPENED ELEGT NRRY TKRDCSTFLR TSETGIVDQV MVTLNQEGYK FCKIRVRSVR IPQIGDKFAS RHGQKG TCG IQYRQEDMPF TCEGITPDII INPHAIPSRM TIGHLIECLQ GKVSANKGEI GDATPFN DA VNVQKISNLL SDYGYHLRGN EVLYNGFTGR KITSQIFIGP TYYQRLKHMV DDKIHSRA R GPIQILNRQP MEGRSRDGGL RFGEMERDCQ IAHGAAQFLR ERLFEASDPY QVHVCNLCG IMAIANTRTH TYECRGCRNK TQISLVRMPY ACKLLFQELM SMSIAPRMMS V

Macromolecule #3: DNA-directed RNA polymerase II subunit RPB3

• Macromolecule: Name: DNA-directed RNA polymerase II subunit RPB3 / type: protein_or_peptide / ID: 3 / Enantiomer: LEVO
• Source (natural): Organism: Sus scrofa (pig)

Sequence

String:

MPYANQPTVR ITELTDENVK FIIENTDLAV ANSIRRVFIA EVPIIAIDWV QIDANSSVLH DEFIAHRLG LIPLTSDDIV DKLQYSRDCT CEEFCPECSV EFTLDVRCNE DQTRHVTSRD L ISNSPRVI PVTSRNRDND PSDYVEQDDI LIVKLRKGQE LRLRAYAKKG FGKEHAKWNP TA GVAFEYD PDNALRHTVY PKPEEWPKSE YSELDEDESQ APYDPNGKPE RFYYNVESCG SLR PETIVL SALSGLKKKL SDLQTQLSHE IQSDVLTIN

Macromolecule #4: RNA polymerase II subunit D

• Macromolecule: Name: RNA polymerase II subunit D / type: protein_or_peptide / ID: 4 / Enantiomer: LEVO
• Source (natural): Organism: Sus scrofa (pig)

Sequence

String:

MWGPAQPYSD SALSPKPRPF RAVFRGSALP FPAVRVEVRG RSMAAGGSDP RAGDVEEDAS QLIFPKEFE TAETLLNSEV HMLLEHRKQQ NESAEDEQEL SEVFMKTLNY TARFSRFKNR E TIASVRSL LLQKKLHKFE LACLANLCPE TAEESKALIP SLEGRFEDEE LQQILDDIQT KR SFQY

Macromolecule #5: RNA polymerase II subunit E

• Macromolecule: Name: RNA polymerase II subunit E / type: protein_or_peptide / ID: 5 / Enantiomer: LEVO
• Source (natural): Organism: Sus scrofa (pig)

Sequence

String:

MDDEEETYRL WKIRKTIMQL CHDRGYLVTQ DELDQTLEEF KAQFGDKPSE GRPRRTDLTV LVAHNDDPT DQMFVFFPEE PKVGIKTIKV YCQRMQEENI TRALIVVQQG MTPSAKQSLV D MAPKYILE QFLQQELLIN ITEHELVPEH VVMTKEEVTE LLARYKLREN QLPRIQAGDP VA RYFGIKR GQVVKIIRPS ETAGRYITYR LVQ

Macromolecule #6: RNA polymerase II subunit F

• Macromolecule: Name: RNA polymerase II subunit F / type: protein_or_peptide / ID: 6 / Enantiomer: LEVO
• Source (natural): Organism: Sus scrofa (pig)

Sequence

String:

MDDEEETYRL WKIRKTIMQL CHDRGYLVTQ DELDQTLEEF KAQFGDKPSE GRPRRTDLTV LVAHNDDPT DQMFVFFPEE PKVGIKTIKV YCQRMQEENI TRALIVVQQG MTPSAKQSLV D MAPKYILE QFLQQELLIN ITEHELVPEH VVMTKEEVTE LLARYKLREN QLPRIQAGDP VA RYFGIKR GQVVKIIRPS ETAGRYITYR LVQ

Macromolecule #7: RNA polymerase II subunit G

• Macromolecule: Name: RNA polymerase II subunit G / type: protein_or_peptide / ID: 7 / Enantiomer: LEVO
• Source (natural): Organism: Sus scrofa (pig)

Sequence

String:

MDDEEETYRL WKIRKTIMQL CHDRGYLVTQ DELDQTLEEF KAQFGDKPSE GRPRRTDLTV LVAHNDDPT DQMFVFFPEE PKVGIKTIKV YCQRMQEENI TRALIVVQQG MTPSAKQSLV D MAPKYILE QFLQQELLIN ITEHELVPEH VVMTKEEVTE LLARYKLREN QLPRIQAGDP VA RYFGIKR GQVVKIIRPS ETAGRYITYR LVQ

Macromolecule #8: 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: 8 / Enantiomer: LEVO
• Source (natural): Organism: Sus scrofa (pig)

Sequence

String:

MAGILFEDIF DVKDIDPEGK KFDRVSRLHC ESESFKMDLI LDVNIQIYPV DLGDKFRLVI ASTLYEDGT LDDGEYNPTD DRPSRADQFE YVMYGKVYRI EGDETSTEAA TRLSAYVSYG G LLMRLQGD ANNLHGFEVD SRVYLLMKKL AF

Macromolecule #9: DNA-directed RNA polymerase II subunit RPB9

• Macromolecule: Name: DNA-directed RNA polymerase II subunit RPB9 / type: protein_or_peptide / ID: 9 / Enantiomer: LEVO
• Source (natural): Organism: Sus scrofa (pig)

Sequence

String:

MEPDGTYEPG FVGIRFCQEC NNMLYPKEDK ENRILLYACR NCDYQQEADN SCIYVNKITH EVDELTQII ADVSQDPTLP RTEDHPCQKC GHKEAVFFQS HSARAEDAMR LYYVCTAPHC G HRWTE

Macromolecule #10: 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: 10 / Enantiomer: LEVO
• Source (natural): Organism: Sus scrofa (pig)

Sequence

String:

MIIPVRCFTC GKIVGNKWEA YLGLLQAEYT EGDALDALGL KRYCCRRMLL AHVDLIEKLL NYAPLEK

Macromolecule #11: DNA-directed RNA polymerase II subunit RPB11-a

• Macromolecule: Name: DNA-directed RNA polymerase II subunit RPB11-a / type: protein_or_peptide / ID: 11 / Enantiomer: LEVO
• Source (natural): Organism: Sus scrofa (pig)

Sequence

String:

MNAPPAFESF LLFEGEKKIT INKDTKVPNA CLFTINKEDH TLGNIIKSQL LKDPQVLFAG YKVPHPLEH KIIIRVQTTP DYSPQEAFTN AITDLISELS LLEERFRVAI KDKQEGIE

Macromolecule #12: RNA polymerase II subunit K

• Macromolecule: Name: RNA polymerase II subunit K / type: protein_or_peptide / ID: 12 / Enantiomer: LEVO
• Source (natural): Organism: Sus scrofa (pig)

Sequence

String:

MDTQKDVQPP KQQPMIYICG ECHTENEIKS RDPIRCRECG YRIMYKKRTK RLVVFDAR

Macromolecule #13: RNA polymerase II elongation factor ELL2

• Macromolecule: Name: RNA polymerase II elongation factor ELL2 / type: protein_or_peptide / ID: 13 / Enantiomer: LEVO
• Source (natural): Organism: Homo sapiens (human)
• Recombinant expression: Organism: Trichoplusia ni (cabbage looper)

Sequence

String:

MAAGGTGGLR EEQRYGLSCG RLGQDNITVL HVKLTETAIR ALETYQSHKN LIPFRPSIQF QGLHGLVKI PKNDPLNEVH NFNFYLSNVG KDNPQGSFDC IQQTFSSSGA SQLNCLGFIQ D KITVCATN DSYQMTRERM TQAEEESRNR STKVIKPGGP YVGKRVQIRK APQAVSDTVP ER KRSTPMN PANTIRKTHS SSTISQRPYR DRVIHLLALK AYKKPELLAR LQKDGVNQKD KNS LGAILQ QVANLNSKDL SYTLKDYVFK ELQRDWPGYS EIDRRSLESV LSRKLNPSQN AAGT SRSES PVCSSRDAVS SPQKRLLDSE FIDPLMNKKA RISHLTNRVP PTLNGHLNPT SEKSA AGLP LPPAAAAIPT PPPLPSTYLP ISHPPQIVNS NSNSPSTPEG RGTQDLPVDS FSQNDS IYE DQQDKYTSRT SLETLPPGSV LLKCPKPMEE NHSMSHKKSK KKSKKHKEKD QIKKHDI ET IEEKEEDLKR EEEIAKLNNS SPNSSGGVKE DCTASMEPSA IELPDYLIKY IAIVSYEQ R QNYKDDFNAE YDEYRALHAR METVARRFIK LDAQRKRLSP GSKEYQNVHE EVLQEYQKI KQSSPNYHEE KYRCEYLHNK LAHIKRLIGE FDQQQAESWS

Macromolecule #14: ELL-associated factor 1

• Macromolecule: Name: ELL-associated factor 1 / type: protein_or_peptide / ID: 14 / Enantiomer: LEVO
• Source (natural): Organism: Homo sapiens (human)
• Recombinant expression: Organism: Trichoplusia ni (cabbage looper)

Sequence

String:

MNGTANPLLD REEHCLRLGE SFEKRPRASF HTIRYDFKPA SIDTSCEGEL QVGKGDEVTI TLPHIPGST PPMTVFKGNK RPYQKDCVLI INHDTGEYVL EKLSSSIQVK KTRAEGSSKI Q ARMEQQPT RPPQTSQPPP PPPPMPFRAP TKPPVGPKTS PLKDNPSPEP QLDDIKRELR AE VDIIEQM SSSSGSSSSD SESSSGSDDD SSSSGGEDNG PASPPQPSHQ QPYNSRPAVA NGT SRPQGS NQLMNTLRND LQLSESGSDS DD

Macromolecule #15: Transcription elongation factor SPT5

• Macromolecule: Name: Transcription elongation factor SPT5 / type: protein_or_peptide / ID: 15 / Enantiomer: LEVO
• Source (natural): Organism: Homo sapiens (human)
• Recombinant expression: Organism: Escherichia coli BL21(DE3) (bacteria)

Sequence

String:

MSDSEDSNFS EEEDSERSSD GEEAEVDEER RSAAGSEKEE EPEDEEEEEE EEEYDEEEEE EDDDRPPKK PRHGGFILDE ADVDDEYEDE DQWEDGAEDI LEKEEIEASN IDNVVLDEDR S GARRLQNL WRDQREEELG EYYMKKYAKS SVGETVYGGS DELSDDITQQ QLLPGVKDPN LW TVKCKIG EERATAISLM RKFIAYQFTD TPLQIKSVVA PEHVKGYIYV EAYKQTHVKQ AIE GVGNLR LGYWNQQMVP IKEMTDVLKV VKEVANLKPK SWVRLKRGIY KDDIAQVDYV EPSQ NTISL KMIPRIDYDR IKARMSLKDW FAKRKKFKRP PQRLFDAEKI RSLGGDVASD GDFLI FEGN RYSRKGFLFK SFAMSAVITE GVKPTLSELE KFEDQPEGID LEVVTESTGK EREHNF QPG DNVEVCEGEL INLQGKILSV DGNKITIMPK HEDLKDMLEF PAQELRKYFK MGDHVKV IA GRFEGDTGLI VRVEENFVIL FSDLTMHELK VLPRDLQLCS ETASGVDVGG QHEWGELV Q LDPQTVGVIV RLERETFQVL NMYGKVVTVR HQAVTRKKDN RFAVALDSEQ NNIHVKDIV KVIDGPHSGR EGEIRHLFRS FAFLHCKKLV ENGGMFVCKT RHLVLAGGSK PRDVTNFTVG GFAPMSPRI SSPMHPSAGG QRGGFGSPGG GSGGMSRGRG RRDNELIGQT VRISQGPYKG Y IGVVKDAT ESTARVELHS TCQTISVDRQ RLTTVGSRRP GGMTSTYGRT PMYGSQTPMY GS GSRTPMY GSQTPLQDGS RTPHYGSQTP LHDGSRTPAQ SGAWDPNNPN TPSRAEEEYE YAF DDEPTP SPQAYGGTPN PQTPGYPDPS SPQVNPQYNP QTPGTPAMYN TDQFSPYAAP SPQG SYQPS PSPQSYHQVA PSPAGYQNTH SPASYHPTPS PMAYQASPSP SPVGYSPMTP GAPSP GGYN PHTPGSGIEQ NSSDWVTTDI QVKVRDTYLD TQVVGQTGVI RSVTGGMCSV YLKDSE KVV SISSEHLEPI TPTKNNKVKV ILGEDREATG VLLSIDGEDG IVRMDLDEQL KILNLRF LG KLLEA

Macromolecule #16: Transcription elongation factor SPT4

• Macromolecule: Name: Transcription elongation factor SPT4 / type: protein_or_peptide / ID: 16 / Enantiomer: LEVO
• Source (natural): Organism: Homo sapiens (human)
• Recombinant expression: Organism: Escherichia coli BL21(DE3) (bacteria)

Sequence

String:

MALETVPKDL RHLRACLLCS LVKTIDQFEY DGCDNCDAYL QMKGNREMVY DCTSSSFDGI IAMMSPEDS WVSKWQRVSN FKPGVYAVSV TGRLPQGIVR ELKSRGVAYK SRDTAIKT

Macromolecule #17: Template DNA

• Macromolecule: Name: Template DNA / type: dna / ID: 17 / Classification: DNA
• Source (natural): Organism: HIV whole-genome vector AA1305#18 (others)

Sequence

String:

GGCAAGCTTT ATTGAGGCTT AAGCAGTGGG TTCAAGGTAC TAGTGTAC

Macromolecule #18: Non-template DNA

• Macromolecule: Name: Non-template DNA / type: dna / ID: 18 / Classification: DNA
• Source (natural): Organism: HIV whole-genome vector AA1305#18 (others)

Sequence

String:

GTACACTAGT ACCTACTCGA GTGACCTTAA GCCTCAATAA AGCTTGCC

Macromolecule #19: RNA

• Macromolecule: Name: RNA / type: rna / ID: 19 ; Details: The last nucleotide (A47) was extended at the 3' of the original synthetic RNA scaffold (46-mer) during complex assembly in the presence of ATP.
• Source (natural): Organism: HIV whole-genome vector AA1305#18 (others)

Sequence

String:

ACCAGAUCUG AGCCUGGGAG CUCUCUGGCU AACUAGGGAA CCCACUA

Experimental details

Structure determination

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

Sample preparation

• Buffer: pH: 7.4
• Vitrification: Cryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 277.15 K / Instrument: FEI VITROBOT MARK IV

Electron microscopy

• Microscope: FEI TITAN KRIOS
• Image recording: Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Detector mode: COUNTING / Average electron dose: 43.21 e/Ų
• Electron beam: Acceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
• Electron optics: Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD
• Experimental equipment: Model: Titan Krios / Image courtesy: FEI Company

Image processing

• Startup model: Type of model: OTHER / Details: Ab initio model created by CryoSPARC
• Final reconstruction: Resolution.type: BY AUTHOR / Resolution: 3.5 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 23136
• Initial angle assignment: Type: MAXIMUM LIKELIHOOD
• Final angle assignment: Type: MAXIMUM LIKELIHOOD

Atomic model buiding 1

• Refinement: Space: REAL