REL 2022-12-07 2023-09-05 2023-09-05 0000-0001-6968-041X Julia Mahamid European Molecular Biology Laboratory Meyerhofstraße 1 Heidelberg Baden-Württemberg Germany 69117 0000-0001-6968-041X Julia Mahamid European Molecular Biology Laboratory Meyerhofstraße 1 Heidelberg Baden-Württemberg Germany 69117 0000-0002-1193-4648 Sara Cuylen-Haering European Molecular Biology Laboratory Meyerhofstraße 1 Heidelberg Baden-Württemberg Germany 69117 Fung HKH Hayashi Y Salo VT Babenko A Zagoriy I Brunner A Ellenberg J Müller CW Cuylen-Haering S Mahamid J German Research Foundation (DFG) 419120233 Germany German Research Foundation (DFG) 402723784 Germany EIPOD fellowship under Marie Sklodowska-Curie Actions COFUND 664726 Germany Human Frontier Science Program (HFSP) CDA00045/2019 France H2020 Marie Curie Actions of the European Commission 101028297 Belgium 174.2 10.6019/EMPIAR-11561 EMDB cell EMD-16303 EMD-18194 Fung HKH Hayashi Y Salo VT Babenko A Zagoriy I Brunner A Ellenberg J Müller CW Cuylen-Haering S Mahamid J Unaligned multi-frame micrographs for dataset 1 (211206) /data/211206/data micrographs - multiframe TIFF TIFF 118 10 1 10 UNSIGNED BYTE 5760 3.425 4092 3.425

Unaligned multi-frame micrographs (frames/*.tif) corresponding to 2 tilt series with corresponding gain reference, defect map and metadata (mdoc/*.mdoc). Data acquired with defocus only, no Volta phase plate, on a K3 BioQuantum detector. See README.md under All Files for more information (also copied below). # Cryo-electron tomography of GEM2-labelled Mito-EGFP in HeLa cells Five independent cryo-ET datasets were collected. Each dataset is organised under the following directory structure, with the directories "data/", "alignment/", and "tomograms/" each designated as an Image Set. - data/ - CountRef.mrc gain reference - defects.tif defect map (K3 data only) - frames/ unaligned, unbinned multi-frame micrographs - mdoc/ MDOC files listing exposure dose, pixel size and tilt angle information for each tilt - alignment/ - TS_###.mrc/ - TS_###.mrc.st sorted, motion-corrected tilt series from WARP - TS_###.mrc.rawtlt tilt angle information corresponding to tilt series - TS_###.mrc.aln AreTomo tilt series alignment solution - TS_###.mrc.xf XF file from AreTomo alignment - mdoc_edited/ MDOC files with entries for discarded tilts during alignment removed - tomograms/ - *.mrc tomograms reconstructed by weighted back-projection in AreTomo, 4-times binned - ######.star combined STAR file listing the refined coordinates of GEM2 particles at 6.85 A/px in this dataset Acquisition parameters differ for each dataset and are specified below. Tilt series were aligned in AreTomo 1.3.1 with the following parameters: 8 by 6 patches (K3 data) or 6 by 6 patches (K2 data), VolZ 2000, AlignZ 1000, TiltAxis ## -1 (as calibrated for the magnification and microsco pe used), TiltCor 0. ## Dataset 1 (211206) - Image Sets 1 to 3 Data acquired with defocus only, no Volta phase plate, on a K3 BioQuantum detector. 2 tilt series. Pixel size, 3.425 A/px. ## Dataset 2 (220330) - Image Sets 4 to 6 Data acquired with defocus only, no Volta phase plate, on a K3 BioQuantum detector. 3 tilt series. Pixel size, 3.425 A/px. ## Dataset 3 (220504) - Image Sets 7 to 9 Data acquired with Volta phase plate on a K2 Summit detector. 1 tilt series. Pixel size, 3.3701 A/px. ## Dataset 4 (220506) - Image Sets 10 to 12 Data acquired with Volta phase plate on a K3 BioQuantum detector. 5 tilt series. Pixel size, 3.425 A/px. ## Dataset 5 (220720) - Image Sets 13 to 15 Data acquired with Volta phase plate on a K3 BioQuantum detector. 6 tilt series. Pixel size, 3.425 A/px.

Tilt series and alignment information for dataset 1 (211206) /data/211206/alignment tilt series MRC MRC 2 1 32 BIT FLOAT 5760 3.425 4092 3.425

Tilt series following motion correction in WARP (*.st), with tilt angle information (*.rawtlt) for tilt series alignment in AreTomo and alignment solutions outputted by AreTomo (*.aln and *.xf). Data acquired with defocus only, no Volta phase plate, on a K3 BioQuantum detector. See README.md under All Files for more information (also copied below). # Cryo-electron tomography of GEM2-labelled Mito-EGFP in HeLa cells Five independent cryo-ET datasets were collected. Each dataset is organised under the following directory structure, with the directories "data/", "alignment/", and "tomograms/" each designated as an Image Set. - data/ - CountRef.mrc gain reference - defects.tif defect map (K3 data only) - frames/ unaligned, unbinned multi-frame micrographs - mdoc/ MDOC files listing exposure dose, pixel size and tilt angle information for each tilt - alignment/ - TS_###.mrc/ - TS_###.mrc.st sorted, motion-corrected tilt series from WARP - TS_###.mrc.rawtlt tilt angle information corresponding to tilt series - TS_###.mrc.aln AreTomo tilt series alignment solution - TS_###.mrc.xf XF file from AreTomo alignment - mdoc_edited/ MDOC files with entries for discarded tilts during alignment removed - tomograms/ - *.mrc tomograms reconstructed by weighted back-projection in AreTomo, 4-times binned - ######.star combined STAR file listing the refined coordinates of GEM2 particles at 6.85 A/px in this dataset Acquisition parameters differ for each dataset and are specified below. Tilt series were aligned in AreTomo 1.3.1 with the following parameters: 8 by 6 patches (K3 data) or 6 by 6 patches (K2 data), VolZ 2000, AlignZ 1000, TiltAxis ## -1 (as calibrated for the magnification and microsco pe used), TiltCor 0. ## Dataset 1 (211206) - Image Sets 1 to 3 Data acquired with defocus only, no Volta phase plate, on a K3 BioQuantum detector. 2 tilt series. Pixel size, 3.425 A/px. ## Dataset 2 (220330) - Image Sets 4 to 6 Data acquired with defocus only, no Volta phase plate, on a K3 BioQuantum detector. 3 tilt series. Pixel size, 3.425 A/px. ## Dataset 3 (220504) - Image Sets 7 to 9 Data acquired with Volta phase plate on a K2 Summit detector. 1 tilt series. Pixel size, 3.3701 A/px. ## Dataset 4 (220506) - Image Sets 10 to 12 Data acquired with Volta phase plate on a K3 BioQuantum detector. 5 tilt series. Pixel size, 3.425 A/px. ## Dataset 5 (220720) - Image Sets 13 to 15 Data acquired with Volta phase plate on a K3 BioQuantum detector. 6 tilt series. Pixel size, 3.425 A/px.

Reconstructed tomograms for dataset 1 (211206) /data/211206/tomograms reconstructed volumes MRC MRC 2 500 32 BIT FLOAT 1022 13.7 1440 13.7

Tomograms (*.mrc) reconstructed by weighted back-projection in AreTomo and 4-times binned, and coordinates from subtomogram averaging of GEM2 particles (*.star). Data acquired with defocus only, no Volta phase plate, on a K3 BioQuantum detector. See README.md under All Files for more information (also copied below). # Cryo-electron tomography of GEM2-labelled Mito-EGFP in HeLa cells Five independent cryo-ET datasets were collected. Each dataset is organised under the following directory structure, with the directories "data/", "alignment/", and "tomograms/" each designated as an Image Set. - data/ - CountRef.mrc gain reference - defects.tif defect map (K3 data only) - frames/ unaligned, unbinned multi-frame micrographs - mdoc/ MDOC files listing exposure dose, pixel size and tilt angle information for each tilt - alignment/ - TS_###.mrc/ - TS_###.mrc.st sorted, motion-corrected tilt series from WARP - TS_###.mrc.rawtlt tilt angle information corresponding to tilt series - TS_###.mrc.aln AreTomo tilt series alignment solution - TS_###.mrc.xf XF file from AreTomo alignment - mdoc_edited/ MDOC files with entries for discarded tilts during alignment removed - tomograms/ - *.mrc tomograms reconstructed by weighted back-projection in AreTomo, 4-times binned - ######.star combined STAR file listing the refined coordinates of GEM2 particles at 6.85 A/px in this dataset Acquisition parameters differ for each dataset and are specified below. Tilt series were aligned in AreTomo 1.3.1 with the following parameters: 8 by 6 patches (K3 data) or 6 by 6 patches (K2 data), VolZ 2000, AlignZ 1000, TiltAxis ## -1 (as calibrated for the magnification and microsco pe used), TiltCor 0. ## Dataset 1 (211206) - Image Sets 1 to 3 Data acquired with defocus only, no Volta phase plate, on a K3 BioQuantum detector. 2 tilt series. Pixel size, 3.425 A/px. ## Dataset 2 (220330) - Image Sets 4 to 6 Data acquired with defocus only, no Volta phase plate, on a K3 BioQuantum detector. 3 tilt series. Pixel size, 3.425 A/px. ## Dataset 3 (220504) - Image Sets 7 to 9 Data acquired with Volta phase plate on a K2 Summit detector. 1 tilt series. Pixel size, 3.3701 A/px. ## Dataset 4 (220506) - Image Sets 10 to 12 Data acquired with Volta phase plate on a K3 BioQuantum detector. 5 tilt series. Pixel size, 3.425 A/px. ## Dataset 5 (220720) - Image Sets 13 to 15 Data acquired with Volta phase plate on a K3 BioQuantum detector. 6 tilt series. Pixel size, 3.425 A/px.

Unaligned multi-frame micrographs for dataset 2 (220330) /data/220330/data micrographs - multiframe TIFF TIFF 178 10 1 10 UNSIGNED BYTE 5760 3.425 4092 3.425

Unaligned multi-frame micrographs (frames/*.tif) corresponding to 3 tilt series with corresponding gain reference, defect map and metadata (mdoc/*.mdoc). Data acquired with defocus only, no Volta phase plate, on a K3 BioQuantum detector. See README.md under 'All Files' for more information (also copied below). # Cryo-electron tomography of GEM2-labelled Mito-EGFP in HeLa cells Five independent cryo-ET datasets were collected. Each dataset is organised under the following directory structure, with the directories "data/", "alignment/", and "tomograms/" each designated as an Image Set. - data/ - CountRef.mrc gain reference - defects.tif defect map (K3 data only) - frames/ unaligned, unbinned multi-frame micrographs - mdoc/ MDOC files listing exposure dose, pixel size and tilt angle information for each tilt - alignment/ - TS_###.mrc/ - TS_###.mrc.st sorted, motion-corrected tilt series from WARP - TS_###.mrc.rawtlt tilt angle information corresponding to tilt series - TS_###.mrc.aln AreTomo tilt series alignment solution - TS_###.mrc.xf XF file from AreTomo alignment - mdoc_edited/ MDOC files with entries for discarded tilts during alignment removed - tomograms/ - *.mrc tomograms reconstructed by weighted back-projection in AreTomo, 4-times binned - ######.star combined STAR file listing the refined coordinates of GEM2 particles at 6.85 A/px in this dataset Acquisition parameters differ for each dataset and are specified below. Tilt series were aligned in AreTomo 1.3.1 with the following parameters: 8 by 6 patches (K3 data) or 6 by 6 patches (K2 data), VolZ 2000, AlignZ 1000, TiltAxis ## -1 (as calibrated for the magnification and microsco pe used), TiltCor 0. ## Dataset 1 (211206) - Image Sets 1 to 3 Data acquired with defocus only, no Volta phase plate, on a K3 BioQuantum detector. 2 tilt series. Pixel size, 3.425 A/px. ## Dataset 2 (220330) - Image Sets 4 to 6 Data acquired with defocus only, no Volta phase plate, on a K3 BioQuantum detector. 3 tilt series. Pixel size, 3.425 A/px. ## Dataset 3 (220504) - Image Sets 7 to 9 Data acquired with Volta phase plate on a K2 Summit detector. 1 tilt series. Pixel size, 3.3701 A/px. ## Dataset 4 (220506) - Image Sets 10 to 12 Data acquired with Volta phase plate on a K3 BioQuantum detector. 5 tilt series. Pixel size, 3.425 A/px. ## Dataset 5 (220720) - Image Sets 13 to 15 Data acquired with Volta phase plate on a K3 BioQuantum detector. 6 tilt series. Pixel size, 3.425 A/px.

Tilt series and alignment information for dataset 2 (220330) /data/220330/alignment tilt series MRC MRC 3 1 32 BIT FLOAT 5760 3.425 4092 3.425

Tilt series following motion correction in WARP (*.st), with tilt angle information (*.rawtlt) for tilt series alignment in AreTomo and alignment solutions outputted by AreTomo (*.aln and *.xf). Data acquired with defocus only, no Volta phase plate, on a K3 BioQuantum detector. See README.md under All Files for more information (also copied below). # Cryo-electron tomography of GEM2-labelled Mito-EGFP in HeLa cells Five independent cryo-ET datasets were collected. Each dataset is organised under the following directory structure, with the directories "data/", "alignment/", and "tomograms/" each designated as an Image Set. - data/ - CountRef.mrc gain reference - defects.tif defect map (K3 data only) - frames/ unaligned, unbinned multi-frame micrographs - mdoc/ MDOC files listing exposure dose, pixel size and tilt angle information for each tilt - alignment/ - TS_###.mrc/ - TS_###.mrc.st sorted, motion-corrected tilt series from WARP - TS_###.mrc.rawtlt tilt angle information corresponding to tilt series - TS_###.mrc.aln AreTomo tilt series alignment solution - TS_###.mrc.xf XF file from AreTomo alignment - mdoc_edited/ MDOC files with entries for discarded tilts during alignment removed - tomograms/ - *.mrc tomograms reconstructed by weighted back-projection in AreTomo, 4-times binned - ######.star combined STAR file listing the refined coordinates of GEM2 particles at 6.85 A/px in this dataset Acquisition parameters differ for each dataset and are specified below. Tilt series were aligned in AreTomo 1.3.1 with the following parameters: 8 by 6 patches (K3 data) or 6 by 6 patches (K2 data), VolZ 2000, AlignZ 1000, TiltAxis ## -1 (as calibrated for the magnification and microsco pe used), TiltCor 0. ## Dataset 1 (211206) - Image Sets 1 to 3 Data acquired with defocus only, no Volta phase plate, on a K3 BioQuantum detector. 2 tilt series. Pixel size, 3.425 A/px. ## Dataset 2 (220330) - Image Sets 4 to 6 Data acquired with defocus only, no Volta phase plate, on a K3 BioQuantum detector. 3 tilt series. Pixel size, 3.425 A/px. ## Dataset 3 (220504) - Image Sets 7 to 9 Data acquired with Volta phase plate on a K2 Summit detector. 1 tilt series. Pixel size, 3.3701 A/px. ## Dataset 4 (220506) - Image Sets 10 to 12 Data acquired with Volta phase plate on a K3 BioQuantum detector. 5 tilt series. Pixel size, 3.425 A/px. ## Dataset 5 (220720) - Image Sets 13 to 15 Data acquired with Volta phase plate on a K3 BioQuantum detector. 6 tilt series. Pixel size, 3.425 A/px.

Reconstructed tomograms for dataset 2 (220330) /data/220330/tomograms reconstructed volumes MRC MRC 3 500 32 BIT FLOAT 1022 13.7 1440 13.7

Tomograms (*.mrc) reconstructed by weighted back-projection in AreTomo and 4-times binned, and coordinates from subtomogram averaging of GEM2 particles (*.star). Data acquired with defocus only, no Volta phase plate, on a K3 BioQuantum detector. See README.md under All Files for more information (also copied below). # Cryo-electron tomography of GEM2-labelled Mito-EGFP in HeLa cells Five independent cryo-ET datasets were collected. Each dataset is organised under the following directory structure, with the directories "data/", "alignment/", and "tomograms/" each designated as an Image Set. - data/ - CountRef.mrc gain reference - defects.tif defect map (K3 data only) - frames/ unaligned, unbinned multi-frame micrographs - mdoc/ MDOC files listing exposure dose, pixel size and tilt angle information for each tilt - alignment/ - TS_###.mrc/ - TS_###.mrc.st sorted, motion-corrected tilt series from WARP - TS_###.mrc.rawtlt tilt angle information corresponding to tilt series - TS_###.mrc.aln AreTomo tilt series alignment solution - TS_###.mrc.xf XF file from AreTomo alignment - mdoc_edited/ MDOC files with entries for discarded tilts during alignment removed - tomograms/ - *.mrc tomograms reconstructed by weighted back-projection in AreTomo, 4-times binned - ######.star combined STAR file listing the refined coordinates of GEM2 particles at 6.85 A/px in this dataset Acquisition parameters differ for each dataset and are specified below. Tilt series were aligned in AreTomo 1.3.1 with the following parameters: 8 by 6 patches (K3 data) or 6 by 6 patches (K2 data), VolZ 2000, AlignZ 1000, TiltAxis ## -1 (as calibrated for the magnification and microsco pe used), TiltCor 0. ## Dataset 1 (211206) - Image Sets 1 to 3 Data acquired with defocus only, no Volta phase plate, on a K3 BioQuantum detector. 2 tilt series. Pixel size, 3.425 A/px. ## Dataset 2 (220330) - Image Sets 4 to 6 Data acquired with defocus only, no Volta phase plate, on a K3 BioQuantum detector. 3 tilt series. Pixel size, 3.425 A/px. ## Dataset 3 (220504) - Image Sets 7 to 9 Data acquired with Volta phase plate on a K2 Summit detector. 1 tilt series. Pixel size, 3.3701 A/px. ## Dataset 4 (220506) - Image Sets 10 to 12 Data acquired with Volta phase plate on a K3 BioQuantum detector. 5 tilt series. Pixel size, 3.425 A/px. ## Dataset 5 (220720) - Image Sets 13 to 15 Data acquired with Volta phase plate on a K3 BioQuantum detector. 6 tilt series. Pixel size, 3.425 A/px.

Unaligned multi-frame micrographs for dataset 3 (220504) /data/220504/data micrographs - multiframe TIFF TIFF 55 8 1 8 UNSIGNED 16 BIT INTEGER 3710 3.3701 3838 3.3701

Unaligned multi-frame micrographs (frames/*.tif) corresponding to 1 tilt series with corresponding gain reference, defect map and metadata (mdoc/*.mdoc). Data acquired with Volta phase plate on a K2 Summit detector. See README.md under All Files for more information (also copied below). # Cryo-electron tomography of GEM2-labelled Mito-EGFP in HeLa cells Five independent cryo-ET datasets were collected. Each dataset is organised under the following directory structure, with the directories "data/", "alignment/", and "tomograms/" each designated as an Image Set. - data/ - CountRef.mrc gain reference - defects.tif defect map (K3 data only) - frames/ unaligned, unbinned multi-frame micrographs - mdoc/ MDOC files listing exposure dose, pixel size and tilt angle information for each tilt - alignment/ - TS_###.mrc/ - TS_###.mrc.st sorted, motion-corrected tilt series from WARP - TS_###.mrc.rawtlt tilt angle information corresponding to tilt series - TS_###.mrc.aln AreTomo tilt series alignment solution - TS_###.mrc.xf XF file from AreTomo alignment - mdoc_edited/ MDOC files with entries for discarded tilts during alignment removed - tomograms/ - *.mrc tomograms reconstructed by weighted back-projection in AreTomo, 4-times binned - ######.star combined STAR file listing the refined coordinates of GEM2 particles at 6.85 A/px in this dataset Acquisition parameters differ for each dataset and are specified below. Tilt series were aligned in AreTomo 1.3.1 with the following parameters: 8 by 6 patches (K3 data) or 6 by 6 patches (K2 data), VolZ 2000, AlignZ 1000, TiltAxis ## -1 (as calibrated for the magnification and microsco pe used), TiltCor 0. ## Dataset 1 (211206) - Image Sets 1 to 3 Data acquired with defocus only, no Volta phase plate, on a K3 BioQuantum detector. 2 tilt series. Pixel size, 3.425 A/px. ## Dataset 2 (220330) - Image Sets 4 to 6 Data acquired with defocus only, no Volta phase plate, on a K3 BioQuantum detector. 3 tilt series. Pixel size, 3.425 A/px. ## Dataset 3 (220504) - Image Sets 7 to 9 Data acquired with Volta phase plate on a K2 Summit detector. 1 tilt series. Pixel size, 3.3701 A/px. ## Dataset 4 (220506) - Image Sets 10 to 12 Data acquired with Volta phase plate on a K3 BioQuantum detector. 5 tilt series. Pixel size, 3.425 A/px. ## Dataset 5 (220720) - Image Sets 13 to 15 Data acquired with Volta phase plate on a K3 BioQuantum detector. 6 tilt series. Pixel size, 3.425 A/px.

Tilt series and alignment information for dataset 3 (220504) /data/220504/alignment tilt series MRC MRC 1 1 32 BIT FLOAT 3710 3.3701 3838 3.3701

Tilt series following motion correction in WARP (*.st), with tilt angle information (*.rawtlt) for tilt series alignment in AreTomo and alignment solutions outputted by AreTomo (*.aln and *.xf). Data acquired with Volta phase plate on a K2 Summit detector. See README.md under All Files for more information (also copied below). # Cryo-electron tomography of GEM2-labelled Mito-EGFP in HeLa cells Five independent cryo-ET datasets were collected. Each dataset is organised under the following directory structure, with the directories "data/", "alignment/", and "tomograms/" each designated as an Image Set. - data/ - CountRef.mrc gain reference - defects.tif defect map (K3 data only) - frames/ unaligned, unbinned multi-frame micrographs - mdoc/ MDOC files listing exposure dose, pixel size and tilt angle information for each tilt - alignment/ - TS_###.mrc/ - TS_###.mrc.st sorted, motion-corrected tilt series from WARP - TS_###.mrc.rawtlt tilt angle information corresponding to tilt series - TS_###.mrc.aln AreTomo tilt series alignment solution - TS_###.mrc.xf XF file from AreTomo alignment - mdoc_edited/ MDOC files with entries for discarded tilts during alignment removed - tomograms/ - *.mrc tomograms reconstructed by weighted back-projection in AreTomo, 4-times binned - ######.star combined STAR file listing the refined coordinates of GEM2 particles at 6.85 A/px in this dataset Acquisition parameters differ for each dataset and are specified below. Tilt series were aligned in AreTomo 1.3.1 with the following parameters: 8 by 6 patches (K3 data) or 6 by 6 patches (K2 data), VolZ 2000, AlignZ 1000, TiltAxis ## -1 (as calibrated for the magnification and microsco pe used), TiltCor 0. ## Dataset 1 (211206) - Image Sets 1 to 3 Data acquired with defocus only, no Volta phase plate, on a K3 BioQuantum detector. 2 tilt series. Pixel size, 3.425 A/px. ## Dataset 2 (220330) - Image Sets 4 to 6 Data acquired with defocus only, no Volta phase plate, on a K3 BioQuantum detector. 3 tilt series. Pixel size, 3.425 A/px. ## Dataset 3 (220504) - Image Sets 7 to 9 Data acquired with Volta phase plate on a K2 Summit detector. 1 tilt series. Pixel size, 3.3701 A/px. ## Dataset 4 (220506) - Image Sets 10 to 12 Data acquired with Volta phase plate on a K3 BioQuantum detector. 5 tilt series. Pixel size, 3.425 A/px. ## Dataset 5 (220720) - Image Sets 13 to 15 Data acquired with Volta phase plate on a K3 BioQuantum detector. 6 tilt series. Pixel size, 3.425 A/px.

Reconstructed tomograms for dataset 3 (220504) /data/220504/tomograms reconstructed volumes MRC MRC 1 500 32 BIT FLOAT 958 13.4804 926 13.4804

Tomograms (*.mrc) reconstructed by weighted back-projection in AreTomo and 4-times binned, and coordinates from subtomogram averaging of GEM2 particles (*.star). Data acquired with Volta phase plate on a K2 Summit detector. See README.md under All Files for more information (also copied below). # Cryo-electron tomography of GEM2-labelled Mito-EGFP in HeLa cells Five independent cryo-ET datasets were collected. Each dataset is organised under the following directory structure, with the directories "data/", "alignment/", and "tomograms/" each designated as an Image Set. - data/ - CountRef.mrc gain reference - defects.tif defect map (K3 data only) - frames/ unaligned, unbinned multi-frame micrographs - mdoc/ MDOC files listing exposure dose, pixel size and tilt angle information for each tilt - alignment/ - TS_###.mrc/ - TS_###.mrc.st sorted, motion-corrected tilt series from WARP - TS_###.mrc.rawtlt tilt angle information corresponding to tilt series - TS_###.mrc.aln AreTomo tilt series alignment solution - TS_###.mrc.xf XF file from AreTomo alignment - mdoc_edited/ MDOC files with entries for discarded tilts during alignment removed - tomograms/ - *.mrc tomograms reconstructed by weighted back-projection in AreTomo, 4-times binned - ######.star combined STAR file listing the refined coordinates of GEM2 particles at 6.85 A/px in this dataset Acquisition parameters differ for each dataset and are specified below. Tilt series were aligned in AreTomo 1.3.1 with the following parameters: 8 by 6 patches (K3 data) or 6 by 6 patches (K2 data), VolZ 2000, AlignZ 1000, TiltAxis ## -1 (as calibrated for the magnification and microsco pe used), TiltCor 0. ## Dataset 1 (211206) - Image Sets 1 to 3 Data acquired with defocus only, no Volta phase plate, on a K3 BioQuantum detector. 2 tilt series. Pixel size, 3.425 A/px. ## Dataset 2 (220330) - Image Sets 4 to 6 Data acquired with defocus only, no Volta phase plate, on a K3 BioQuantum detector. 3 tilt series. Pixel size, 3.425 A/px. ## Dataset 3 (220504) - Image Sets 7 to 9 Data acquired with Volta phase plate on a K2 Summit detector. 1 tilt series. Pixel size, 3.3701 A/px. ## Dataset 4 (220506) - Image Sets 10 to 12 Data acquired with Volta phase plate on a K3 BioQuantum detector. 5 tilt series. Pixel size, 3.425 A/px. ## Dataset 5 (220720) - Image Sets 13 to 15 Data acquired with Volta phase plate on a K3 BioQuantum detector. 6 tilt series. Pixel size, 3.425 A/px.

Unaligned multi-frame micrographs for dataset 4 (220506) /data/220506/data micrographs - multiframe TIFF TIFF 293 10 1 10 UNSIGNED BYTE 5760 3.425 4092 3.425

Unaligned multi-frame micrographs (frames/*.tif) corresponding to 5 tilt series with corresponding gain reference, defect map and metadata (mdoc/*.mdoc). Data acquired with Volta phase plate on a K3 BioQuantum detector. See README.md under All Files for more information (also copied below). # Cryo-electron tomography of GEM2-labelled Mito-EGFP in HeLa cells Five independent cryo-ET datasets were collected. Each dataset is organised under the following directory structure, with the directories "data/", "alignment/", and "tomograms/" each designated as an Image Set. - data/ - CountRef.mrc gain reference - defects.tif defect map (K3 data only) - frames/ unaligned, unbinned multi-frame micrographs - mdoc/ MDOC files listing exposure dose, pixel size and tilt angle information for each tilt - alignment/ - TS_###.mrc/ - TS_###.mrc.st sorted, motion-corrected tilt series from WARP - TS_###.mrc.rawtlt tilt angle information corresponding to tilt series - TS_###.mrc.aln AreTomo tilt series alignment solution - TS_###.mrc.xf XF file from AreTomo alignment - mdoc_edited/ MDOC files with entries for discarded tilts during alignment removed - tomograms/ - *.mrc tomograms reconstructed by weighted back-projection in AreTomo, 4-times binned - ######.star combined STAR file listing the refined coordinates of GEM2 particles at 6.85 A/px in this dataset Acquisition parameters differ for each dataset and are specified below. Tilt series were aligned in AreTomo 1.3.1 with the following parameters: 8 by 6 patches (K3 data) or 6 by 6 patches (K2 data), VolZ 2000, AlignZ 1000, TiltAxis ## -1 (as calibrated for the magnification and microsco pe used), TiltCor 0. ## Dataset 1 (211206) - Image Sets 1 to 3 Data acquired with defocus only, no Volta phase plate, on a K3 BioQuantum detector. 2 tilt series. Pixel size, 3.425 A/px. ## Dataset 2 (220330) - Image Sets 4 to 6 Data acquired with defocus only, no Volta phase plate, on a K3 BioQuantum detector. 3 tilt series. Pixel size, 3.425 A/px. ## Dataset 3 (220504) - Image Sets 7 to 9 Data acquired with Volta phase plate on a K2 Summit detector. 1 tilt series. Pixel size, 3.3701 A/px. ## Dataset 4 (220506) - Image Sets 10 to 12 Data acquired with Volta phase plate on a K3 BioQuantum detector. 5 tilt series. Pixel size, 3.425 A/px. ## Dataset 5 (220720) - Image Sets 13 to 15 Data acquired with Volta phase plate on a K3 BioQuantum detector. 6 tilt series. Pixel size, 3.425 A/px.

Tilt series and alignment information for dataset 4 (220506) /data/220506/alignment tilt series MRC MRC 5 1 32 BIT FLOAT 5760 3.425 4092 3.425

Tilt series following motion correction in WARP (*.st), with tilt angle information (*.rawtlt) for tilt series alignment in AreTomo and alignment solutions outputted by AreTomo (*.aln and *.xf). Data acquired with Volta phase plate on a K3 BioQuantum detector. See README.md under All Files for more information (also copied below). # Cryo-electron tomography of GEM2-labelled Mito-EGFP in HeLa cells Five independent cryo-ET datasets were collected. Each dataset is organised under the following directory structure, with the directories "data/", "alignment/", and "tomograms/" each designated as an Image Set. - data/ - CountRef.mrc gain reference - defects.tif defect map (K3 data only) - frames/ unaligned, unbinned multi-frame micrographs - mdoc/ MDOC files listing exposure dose, pixel size and tilt angle information for each tilt - alignment/ - TS_###.mrc/ - TS_###.mrc.st sorted, motion-corrected tilt series from WARP - TS_###.mrc.rawtlt tilt angle information corresponding to tilt series - TS_###.mrc.aln AreTomo tilt series alignment solution - TS_###.mrc.xf XF file from AreTomo alignment - mdoc_edited/ MDOC files with entries for discarded tilts during alignment removed - tomograms/ - *.mrc tomograms reconstructed by weighted back-projection in AreTomo, 4-times binned - ######.star combined STAR file listing the refined coordinates of GEM2 particles at 6.85 A/px in this dataset Acquisition parameters differ for each dataset and are specified below. Tilt series were aligned in AreTomo 1.3.1 with the following parameters: 8 by 6 patches (K3 data) or 6 by 6 patches (K2 data), VolZ 2000, AlignZ 1000, TiltAxis ## -1 (as calibrated for the magnification and microsco pe used), TiltCor 0. ## Dataset 1 (211206) - Image Sets 1 to 3 Data acquired with defocus only, no Volta phase plate, on a K3 BioQuantum detector. 2 tilt series. Pixel size, 3.425 A/px. ## Dataset 2 (220330) - Image Sets 4 to 6 Data acquired with defocus only, no Volta phase plate, on a K3 BioQuantum detector. 3 tilt series. Pixel size, 3.425 A/px. ## Dataset 3 (220504) - Image Sets 7 to 9 Data acquired with Volta phase plate on a K2 Summit detector. 1 tilt series. Pixel size, 3.3701 A/px. ## Dataset 4 (220506) - Image Sets 10 to 12 Data acquired with Volta phase plate on a K3 BioQuantum detector. 5 tilt series. Pixel size, 3.425 A/px. ## Dataset 5 (220720) - Image Sets 13 to 15 Data acquired with Volta phase plate on a K3 BioQuantum detector. 6 tilt series. Pixel size, 3.425 A/px.

Reconstructed tomograms for dataset 4 (220506) /data/220506/tomograms reconstructed volumes MRC MRC 5 500 32 BIT FLOAT 1022 13.7 1440 13.7

Tomograms (*.mrc) reconstructed by weighted back-projection in AreTomo and 4-times binned, and coordinates from subtomogram averaging of GEM2 particles (*.star). Data acquired with Volta phase plate on a K3 BioQuantum detector. See README.md under All Files for more information (also copied below). # Cryo-electron tomography of GEM2-labelled Mito-EGFP in HeLa cells Five independent cryo-ET datasets were collected. Each dataset is organised under the following directory structure, with the directories "data/", "alignment/", and "tomograms/" each designated as an Image Set. - data/ - CountRef.mrc gain reference - defects.tif defect map (K3 data only) - frames/ unaligned, unbinned multi-frame micrographs - mdoc/ MDOC files listing exposure dose, pixel size and tilt angle information for each tilt - alignment/ - TS_###.mrc/ - TS_###.mrc.st sorted, motion-corrected tilt series from WARP - TS_###.mrc.rawtlt tilt angle information corresponding to tilt series - TS_###.mrc.aln AreTomo tilt series alignment solution - TS_###.mrc.xf XF file from AreTomo alignment - mdoc_edited/ MDOC files with entries for discarded tilts during alignment removed - tomograms/ - *.mrc tomograms reconstructed by weighted back-projection in AreTomo, 4-times binned - ######.star combined STAR file listing the refined coordinates of GEM2 particles at 6.85 A/px in this dataset Acquisition parameters differ for each dataset and are specified below. Tilt series were aligned in AreTomo 1.3.1 with the following parameters: 8 by 6 patches (K3 data) or 6 by 6 patches (K2 data), VolZ 2000, AlignZ 1000, TiltAxis ## -1 (as calibrated for the magnification and microsco pe used), TiltCor 0. ## Dataset 1 (211206) - Image Sets 1 to 3 Data acquired with defocus only, no Volta phase plate, on a K3 BioQuantum detector. 2 tilt series. Pixel size, 3.425 A/px. ## Dataset 2 (220330) - Image Sets 4 to 6 Data acquired with defocus only, no Volta phase plate, on a K3 BioQuantum detector. 3 tilt series. Pixel size, 3.425 A/px. ## Dataset 3 (220504) - Image Sets 7 to 9 Data acquired with Volta phase plate on a K2 Summit detector. 1 tilt series. Pixel size, 3.3701 A/px. ## Dataset 4 (220506) - Image Sets 10 to 12 Data acquired with Volta phase plate on a K3 BioQuantum detector. 5 tilt series. Pixel size, 3.425 A/px. ## Dataset 5 (220720) - Image Sets 13 to 15 Data acquired with Volta phase plate on a K3 BioQuantum detector. 6 tilt series. Pixel size, 3.425 A/px.

Unaligned multi-frame micrographs for dataset 5 (220720) /data/220720/data micrographs - multiframe TIFF TIFF 340 10 1 10 UNSIGNED BYTE 5760 3.425 4092 3.425

Unaligned multi-frame micrographs (frames/*.tif) corresponding to 6 tilt series with corresponding gain reference, defect map and metadata (mdoc/*.mdoc). Data acquired with Volta phase plate on a K3 BioQuantum detector. See README.md under All Files for more information (also copied below). # Cryo-electron tomography of GEM2-labelled Mito-EGFP in HeLa cells Five independent cryo-ET datasets were collected. Each dataset is organised under the following directory structure, with the directories "data/", "alignment/", and "tomograms/" each designated as an Image Set. - data/ - CountRef.mrc gain reference - defects.tif defect map (K3 data only) - frames/ unaligned, unbinned multi-frame micrographs - mdoc/ MDOC files listing exposure dose, pixel size and tilt angle information for each tilt - alignment/ - TS_###.mrc/ - TS_###.mrc.st sorted, motion-corrected tilt series from WARP - TS_###.mrc.rawtlt tilt angle information corresponding to tilt series - TS_###.mrc.aln AreTomo tilt series alignment solution - TS_###.mrc.xf XF file from AreTomo alignment - mdoc_edited/ MDOC files with entries for discarded tilts during alignment removed - tomograms/ - *.mrc tomograms reconstructed by weighted back-projection in AreTomo, 4-times binned - ######.star combined STAR file listing the refined coordinates of GEM2 particles at 6.85 A/px in this dataset Acquisition parameters differ for each dataset and are specified below. Tilt series were aligned in AreTomo 1.3.1 with the following parameters: 8 by 6 patches (K3 data) or 6 by 6 patches (K2 data), VolZ 2000, AlignZ 1000, TiltAxis ## -1 (as calibrated for the magnification and microsco pe used), TiltCor 0. ## Dataset 1 (211206) - Image Sets 1 to 3 Data acquired with defocus only, no Volta phase plate, on a K3 BioQuantum detector. 2 tilt series. Pixel size, 3.425 A/px. ## Dataset 2 (220330) - Image Sets 4 to 6 Data acquired with defocus only, no Volta phase plate, on a K3 BioQuantum detector. 3 tilt series. Pixel size, 3.425 A/px. ## Dataset 3 (220504) - Image Sets 7 to 9 Data acquired with Volta phase plate on a K2 Summit detector. 1 tilt series. Pixel size, 3.3701 A/px. ## Dataset 4 (220506) - Image Sets 10 to 12 Data acquired with Volta phase plate on a K3 BioQuantum detector. 5 tilt series. Pixel size, 3.425 A/px. ## Dataset 5 (220720) - Image Sets 13 to 15 Data acquired with Volta phase plate on a K3 BioQuantum detector. 6 tilt series. Pixel size, 3.425 A/px.

Tilt series and alignment information for dataset 5 (220720) /data/220720/alignment tilt series MRC MRC 6 1 32 BIT FLOAT 5760 3.425 4092 3.425

Tilt series following motion correction in WARP (*.st), with tilt angle information (*.rawtlt) for tilt series alignment in AreTomo and alignment solutions outputted by AreTomo (*.aln and *.xf). Data acquired with Volta phase plate on a K3 BioQuantum detector. See README.md under All Files for more information (also copied below). # Cryo-electron tomography of GEM2-labelled Mito-EGFP in HeLa cells Five independent cryo-ET datasets were collected. Each dataset is organised under the following directory structure, with the directories "data/", "alignment/", and "tomograms/" each designated as an Image Set. - data/ - CountRef.mrc gain reference - defects.tif defect map (K3 data only) - frames/ unaligned, unbinned multi-frame micrographs - mdoc/ MDOC files listing exposure dose, pixel size and tilt angle information for each tilt - alignment/ - TS_###.mrc/ - TS_###.mrc.st sorted, motion-corrected tilt series from WARP - TS_###.mrc.rawtlt tilt angle information corresponding to tilt series - TS_###.mrc.aln AreTomo tilt series alignment solution - TS_###.mrc.xf XF file from AreTomo alignment - mdoc_edited/ MDOC files with entries for discarded tilts during alignment removed - tomograms/ - *.mrc tomograms reconstructed by weighted back-projection in AreTomo, 4-times binned - ######.star combined STAR file listing the refined coordinates of GEM2 particles at 6.85 A/px in this dataset Acquisition parameters differ for each dataset and are specified below. Tilt series were aligned in AreTomo 1.3.1 with the following parameters: 8 by 6 patches (K3 data) or 6 by 6 patches (K2 data), VolZ 2000, AlignZ 1000, TiltAxis ## -1 (as calibrated for the magnification and microsco pe used), TiltCor 0. ## Dataset 1 (211206) - Image Sets 1 to 3 Data acquired with defocus only, no Volta phase plate, on a K3 BioQuantum detector. 2 tilt series. Pixel size, 3.425 A/px. ## Dataset 2 (220330) - Image Sets 4 to 6 Data acquired with defocus only, no Volta phase plate, on a K3 BioQuantum detector. 3 tilt series. Pixel size, 3.425 A/px. ## Dataset 3 (220504) - Image Sets 7 to 9 Data acquired with Volta phase plate on a K2 Summit detector. 1 tilt series. Pixel size, 3.3701 A/px. ## Dataset 4 (220506) - Image Sets 10 to 12 Data acquired with Volta phase plate on a K3 BioQuantum detector. 5 tilt series. Pixel size, 3.425 A/px. ## Dataset 5 (220720) - Image Sets 13 to 15 Data acquired with Volta phase plate on a K3 BioQuantum detector. 6 tilt series. Pixel size, 3.425 A/px.

Reconstructed tomograms for dataset 5 (220720) /data/220720/tomograms reconstructed volumes MRC MRC 6 500 32 BIT FLOAT 1022 13.7 1440 13.7

Tomograms (*.mrc) reconstructed by weighted back-projection in AreTomo and 4-times binned, and coordinates from subtomogram averaging of GEM2 particles (*.star). Data acquired with Volta phase plate on a K3 BioQuantum detector. See README.md under All Files for more information (also copied below). # Cryo-electron tomography of GEM2-labelled Mito-EGFP in HeLa cells Five independent cryo-ET datasets were collected. Each dataset is organised under the following directory structure, with the directories "data/", "alignment/", and "tomograms/" each designated as an Image Set. - data/ - CountRef.mrc gain reference - defects.tif defect map (K3 data only) - frames/ unaligned, unbinned multi-frame micrographs - mdoc/ MDOC files listing exposure dose, pixel size and tilt angle information for each tilt - alignment/ - TS_###.mrc/ - TS_###.mrc.st sorted, motion-corrected tilt series from WARP - TS_###.mrc.rawtlt tilt angle information corresponding to tilt series - TS_###.mrc.aln AreTomo tilt series alignment solution - TS_###.mrc.xf XF file from AreTomo alignment - mdoc_edited/ MDOC files with entries for discarded tilts during alignment removed - tomograms/ - *.mrc tomograms reconstructed by weighted back-projection in AreTomo, 4-times binned - ######.star combined STAR file listing the refined coordinates of GEM2 particles at 6.85 A/px in this dataset Acquisition parameters differ for each dataset and are specified below. Tilt series were aligned in AreTomo 1.3.1 with the following parameters: 8 by 6 patches (K3 data) or 6 by 6 patches (K2 data), VolZ 2000, AlignZ 1000, TiltAxis ## -1 (as calibrated for the magnification and microsco pe used), TiltCor 0. ## Dataset 1 (211206) - Image Sets 1 to 3 Data acquired with defocus only, no Volta phase plate, on a K3 BioQuantum detector. 2 tilt series. Pixel size, 3.425 A/px. ## Dataset 2 (220330) - Image Sets 4 to 6 Data acquired with defocus only, no Volta phase plate, on a K3 BioQuantum detector. 3 tilt series. Pixel size, 3.425 A/px. ## Dataset 3 (220504) - Image Sets 7 to 9 Data acquired with Volta phase plate on a K2 Summit detector. 1 tilt series. Pixel size, 3.3701 A/px. ## Dataset 4 (220506) - Image Sets 10 to 12 Data acquired with Volta phase plate on a K3 BioQuantum detector. 5 tilt series. Pixel size, 3.425 A/px. ## Dataset 5 (220720) - Image Sets 13 to 15 Data acquired with Volta phase plate on a K3 BioQuantum detector. 6 tilt series. Pixel size, 3.425 A/px.