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- EMDB-16443: Cryo-EM structure of yeast mitochondrial RNA polymerase transcrip... -
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Open data
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Basic information
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Title | Cryo-EM structure of yeast mitochondrial RNA polymerase transcription initiation complex with 8-mer RNA, pppGpGpUpApApApUpG (IC8) | |||||||||
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![]() | gene transcription / polymerase / RDRP / MTF1 / RPO41 / POLRMT / mtRNAP / DNA / transcription initiation / RNA polymerase / mitochondria / IC8 / TRANSCRIPTION | |||||||||
Function / homology | ![]() Mitochondrial transcription initiation / mitochondrial DNA-directed RNA polymerase complex / mitochondrial transcription factor activity / transcription initiation at mitochondrial promoter / mitochondrial promoter sequence-specific DNA binding / mitochondrial transcription / mitochondrial genome maintenance / DNA primase activity / DNA replication, synthesis of primer / positive regulation of DNA-templated transcription, elongation ...Mitochondrial transcription initiation / mitochondrial DNA-directed RNA polymerase complex / mitochondrial transcription factor activity / transcription initiation at mitochondrial promoter / mitochondrial promoter sequence-specific DNA binding / mitochondrial transcription / mitochondrial genome maintenance / DNA primase activity / DNA replication, synthesis of primer / positive regulation of DNA-templated transcription, elongation / mitochondrial nucleoid / Transferases; Transferring one-carbon groups; Methyltransferases / methyltransferase activity / mitochondrial intermembrane space / DNA-directed 5'-3' RNA polymerase activity / DNA-directed RNA polymerase / methylation / mitochondrial matrix / mitochondrion / DNA binding / RNA binding / cytoplasm Similarity search - Function | |||||||||
Biological species | ![]() ![]() | |||||||||
Method | single particle reconstruction / cryo EM / Resolution: 3.62 Å | |||||||||
![]() | Goovaerts Q / Shen J / Patel SS / Das K | |||||||||
Funding support | ![]()
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![]() | ![]() Title: Structures illustrate step-by-step mitochondrial transcription initiation. Authors: Quinten Goovaerts / Jiayu Shen / Brent De Wijngaert / Urmimala Basu / Smita S Patel / Kalyan Das / ![]() ![]() Abstract: Transcription initiation is a key regulatory step in gene expression during which RNA polymerase (RNAP) initiates RNA synthesis de novo, and the synthesized RNA at a specific length triggers the ...Transcription initiation is a key regulatory step in gene expression during which RNA polymerase (RNAP) initiates RNA synthesis de novo, and the synthesized RNA at a specific length triggers the transition to the elongation phase. Mitochondria recruit a single-subunit RNAP and one or two auxiliary factors to initiate transcription. Previous studies have revealed the molecular architectures of yeast and human mitochondrial RNAP initiation complexes (ICs). Here we provide a comprehensive, stepwise mechanism of transcription initiation by solving high-resolution cryogenic electron microscopy (cryo-EM) structures of yeast mitochondrial RNAP and the transcription factor Mtf1 catalysing two- to eight-nucleotide RNA synthesis at single-nucleotide addition steps. The growing RNA-DNA is accommodated in the polymerase cleft by template scrunching and non-template reorganization, creating stressed intermediates. During early initiation, non-template strand scrunching and unscrunching destabilize the short two- and three-nucleotide RNAs, triggering abortive synthesis. Subsequently, the non-template reorganizes into a base-stacked staircase-like structure supporting processive five- to eight-nucleotide RNA synthesis. The expanded non-template staircase and highly scrunched template in IC8 destabilize the promoter interactions with Mtf1 to facilitate initiation bubble collapse and promoter escape for the transition from initiation to the elongation complex (EC). The series of transcription initiation steps, each guided by the interplay of multiple structural components, reveal a finely tuned mechanism for potential regulatory control. | |||||||||
History |
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Structure visualization
Supplemental images |
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Downloads & links
-EMDB archive
Map data | ![]() | 7.8 MB | ![]() | |
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Header (meta data) | ![]() ![]() | 28.5 KB 28.5 KB | Display Display | ![]() |
FSC (resolution estimation) | ![]() | 8 KB | Display | ![]() |
Images | ![]() | 49.9 KB | ||
Masks | ![]() | 8.4 MB | ![]() | |
Filedesc metadata | ![]() | 8 KB | ||
Others | ![]() ![]() | 7.8 MB 7.8 MB | ||
Archive directory | ![]() ![]() | HTTPS FTP |
-Related structure data
Related structure data | ![]() 8c5uMC ![]() 8ap1C ![]() 8attC ![]() 8atvC ![]() 8atwC ![]() 8c5sC ![]() 8q63C C: citing same article ( M: atomic model generated by this map |
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Similar structure data | Similarity search - Function & homology ![]() |
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Links
EMDB pages | ![]() ![]() |
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Related items in Molecule of the Month |
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Map
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Voxel size | X=Y=Z: 0.97 Å | ||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||
Details | EMDB XML:
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-Supplemental data
-Mask #1
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-Half map: #2
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Density Histograms |
-Half map: #1
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Density Histograms |
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Sample components
+Entire : yeast mitochondrial RNA polymerase transcription initiation compl...
+Supramolecule #1: yeast mitochondrial RNA polymerase transcription initiation compl...
+Supramolecule #2: DNA-directed RNA polymerase, mitochondrial
+Supramolecule #3: DNA (37-mer)
+Supramolecule #4: Mitochondrial transcription factor 1
+Supramolecule #5: RNA (pppGpGpUpApApApUpG)
+Macromolecule #1: Mitochondrial transcription factor 1
+Macromolecule #2: DNA-directed RNA polymerase, mitochondrial
+Macromolecule #3: Non-template DNA (37-MER)
+Macromolecule #4: Template DNA (37-MER)
+Macromolecule #5: RNA (pppGpGpUpApApApUpG)
+Macromolecule #6: GUANOSINE-5'-TRIPHOSPHATE
-Experimental details
-Structure determination
Method | cryo EM |
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![]() | single particle reconstruction |
Aggregation state | particle |
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Sample preparation
Concentration | 0.5 mg/mL |
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Buffer | pH: 7 Details: 50 mM Bis-Tris propane; 100 mM NaCl, 5 mM MgCl2, 1 mM EDTA, 2 mM DTT |
Grid | Model: Quantifoil R1.2/1.3 / Material: GOLD / Mesh: 300 / Support film - Material: CARBON / Support film - topology: HOLEY / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 45 sec. / Pretreatment - Atmosphere: AIR / Pretreatment - Pressure: 0.03 kPa |
Vitrification | Cryogen name: ETHANE / Chamber humidity: 95 % / Chamber temperature: 281.15 K / Instrument: LEICA EM GP / Details: 3 ul sample, back blotting for 12 seconds. |
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Electron microscopy
Microscope | TFS GLACIOS |
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Image recording | Film or detector model: FEI FALCON III (4k x 4k) / Detector mode: COUNTING / Number grids imaged: 1 / Number real images: 1409 / Average exposure time: 37.06 sec. / Average electron dose: 40.0 e/Å2 |
Electron beam | Acceleration voltage: 200 kV / Electron source: ![]() |
Electron optics | C2 aperture diameter: 50.0 µm / Calibrated defocus max: 2.6 µm / Calibrated defocus min: 0.55 µm / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: 1.8 µm / Nominal defocus min: 0.8 µm / Nominal magnification: 150000 |
Sample stage | Specimen holder model: OTHER / Cooling holder cryogen: NITROGEN |
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Image processing
-Atomic model buiding 1
Details | Realspace refinement |
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Refinement | Space: REAL / Protocol: FLEXIBLE FIT / Overall B value: 80 / Target criteria: Correlation coefficient |
Output model | ![]() PDB-8c5u: |