Dataset Information

The ribotoxic stress response drives UV-mediated cell death.

ABSTRACT: Supporting raw MS data for paper (doi: 10.1016/j.cell.2024.05.018) by Sinha N.K. et al, titled "The ribotoxic stress response drives UV-mediated cell death". Index of MS files and labels Table S1 - HaCaT Timecourse. 1-1 - Proteome (AO3604-3627 | Unimod:35; 2016; 4 | 126 -UT_01; 127n - UT_02; 127c - UT_03; 128n -1min_01; 128c - 1min_02; 129n - 1min_03; 129c - 1min_04; 130n - 5min_01; 130c - 5min_02; 131n - 5min_03; 131c - 15min_01; 132n - 15min_02; 132c - 15min_03; 133n - 30min_01; 133c - 30min_02; 134n - 30min_03) 1-2 - Phospho (AO3628-3651 | Unimod:35; 2016; 4; 21 | 126 -UT_01; 127n - UT_02; 127c - UT_03; 128n -1min_01; 128c - 1min_02; 129n - 1min_03; 129c - 1min_04; 130n - 5min_01; 130c - 5min_02; 131n - 5min_03; 131c - 15min_01; 132n - 15min_02; 132c - 15min_03; 133n - 30min_01; 133c - 30min_02; 134n - 30min_03) Table S2 - MCF10a KO and inhibitor (Set1 - ZAK & Set 2 - GCN2). 2-1-1 - Proteome (Set 1 - AO4019-4042 | Unimod:35; 2016; 4 | 126 - WT_UT_DMSO01; 127c - WT_UT_DMSO02; 128n - WT_UT_ZAKi01; 128c - WT_UT_ZAKi02; 129n - WT_UV_DMSO01; 129c -WT_UV_DMSO02; 130n - WT_UV_ZAKi01; 130c - WT_UV_ZAKi02; 131n - ZAK-KO_UT_DMSO01; 131c - ZAK-KO_UT_DMSO02; 132n - ZAK-KO_UT_ZAKi01; 132c - ZAK-KO_UT_ZAKi02; 133n - ZAK-KO_UV_DMSO01; 133c - ZAK-KO_UV_DMSO02; 134n - ZAK-KO_UV_ZAKi01; 134c -ZAK-KO_UV_ZAKi02) 2-1-2 - Proteome (Set 2 - AO4049-4072 | Unimod:35; 2016; 4 | 126 - WT_UT_DMSO01; 127c - WT_UT_DMSO02; 128n - WT_UT_GCN2i01; 128c - WT_UT_GCN2i02; 129n - WT_UV_DMSO01; 129c -WT_UV_DMSO02; 130n - WT_UV_GCN2i01; 130c - WT_UV_GCN2i02; 131n - GCN2-KO_UT_DMSO01; 131c - GCN2-KO_UT_DMSO02; 132n - GCN2-KO_UT_GCN2i01; 132c - GCN2-KO_UT_GCN2i02; 133n - GCN2-KO_UV_DMSO01; 133c - GCN2-KO_UV_DMSO02; 134n - GCN2-KO_UV_GCN2i01; 135n -GCN2-KO_UV_GCN2i02) 2-2-1 - Phospho (Set 1 - AO4013-4019 | Unimod:35; 2016; 4; 21 | 126 - WT_UT_DMSO01; 127c - WT_UT_DMSO02; 128n - WT_UT_ZAKi01; 128c - WT_UT_ZAKi02; 129n - WT_UV_DMSO01; 129c -WT_UV_DMSO02; 130n - WT_UV_ZAKi01; 130c - WT_UV_ZAKi02; 131n - ZAK-KO_UT_DMSO01; 131c - ZAK-KO_UT_DMSO02; 132n - ZAK-KO_UT_ZAKi01; 132c - ZAK-KO_UT_ZAKi02; 133n - ZAK-KO_UV_DMSO01; 133c - ZAK-KO_UV_DMSO02; 134n - ZAK-KO_UV_ZAKi01; 134c -ZAK-KO_UV_ZAKi02) 2-2-2 - Phospho (Set 2 - AO4043-4048 | Unimod:35; 2016; 4; 21 | 126 - WT_UT_DMSO01; 127c - WT_UT_DMSO02; 128n - WT_UT_GCN2i01; 128c - WT_UT_GCN2i02; 129n - WT_UV_DMSO01; 129c -WT_UV_DMSO02; 130n - WT_UV_GCN2i01; 130c - WT_UV_GCN2i02; 131n - GCN2-KO_UT_DMSO01; 131c - GCN2-KO_UT_DMSO02; 132n - GCN2-KO_UT_GCN2i01; 132c - GCN2-KO_UT_GCN2i02; 133n - GCN2-KO_UV_DMSO01; 133c - GCN2-KO_UV_DMSO02; 134n - GCN2-KO_UV_GCN2i01; 135n -GCN2-KO_UV_GCN2i02) Table S3 - HaCaT Kinase inhibitors (Set 1 & Set 2). 3-1-1 - Proteome (Set 1 - AO3766-3777 | Unimod:35; 2016; 4 | 126 - DMSO_UT01; 127n - DMSO_UT02; 127c - DMSO_UV01; 128n - DMSO_UV02; 128c - ZAKi(Nil)_UT01; 129n - ZAKi(Nil)_UT02; 129c - ZAKi(Nil)_UV01; 130n - ZAKi(Nil)_UV02; 130c - p38i_UT01; 131n - p38i_UT02; 131c - p38i_UV01; 132n - p38i_UV02; 132c - JNKi_UT01; 133n - JNKi_UT02; 133c - JNKi_UV01; 134n - JNKi_UV02) 3-1-2 - Proteome (Set 2 - AO3778-3789 | Unimod:35; 2016; 4 | 126 - DMSO_UT01; 127n - DMSO_UT02; 127c - DMSO_UV01; 128n - DMSO_UV02; 128c - ZAKi(Vem)_UT01; 129n - ZAKi(Vem)_UT02; 129c - ZAKi(Vem)_UV01; 130n - ZAKi(Vem)_UV02; 130c - GCN2i_UT01; 131n - GCN2i_UT02; 131c - GCN2i_UV01; 132n - GCN2i_UV02; 132c - ATRi_UT01; 133n - ATRi_UT02; 133c - ATRi_UV01; 134n - ATRi_UV02) 3-2-1 - Phospho (Set 1 - AO3754-3759 | Unimod:35; 2016; 4; 21 | 126 - DMSO_UT01; 127n - DMSO_UT02; 127c - DMSO_UV01; 128n - DMSO_UV02; 128c - ZAKi(Nil)_UT01; 129n - ZAKi(Nil)_UT02; 129c - ZAKi(Nil)_UV01; 130n - ZAKi(Nil)_UV02; 130c - p38i_UT01; 131n - p38i_UT02; 131c - p38i_UV01; 132n - p38i_UV02; 132c - JNKi_UT01; 133n - JNKi_UT02; 133c - JNKi_UV01; 134n - JNKi_UV02) 3-2-2 - Phospho (Set 2 - AO3760-3765 | Unimod:35; 2016; 4; 21 | 126 - DMSO_UT01; 127n - DMSO_UT02; 127c - DMSO_UV01; 128n - DMSO_UV02; 128c - ZAKi(Vem)_UT01; 129n - ZAKi(Vem)_UT02; 129c - ZAKi(Vem)_UV01; 130n - ZAKi(Vem)_UV02; 130c - GCN2i_UT01; 131n - GCN2i_UT02; 131c - GCN2i_UV01; 132n - GCN2i_UV02; 132c - ATRi_UT01; 133n - ATRi_UT02; 133c - ATRi_UV01; 134n - ATRi_UV02). Tabel S4 - ZAK affinity purification. 4-1 - Label-free ZAK pulldown (mNG-ZAK-WT labeled files | Unimod:35; 4| Untreated or ANS treated cells) Additional file: AO4072_GCN2_RSP10_tMS2: Targeted RPS10-Kgg search | Unimod:35; 2016; 4; 492 | TMT label as 2-1-2.

INSTRUMENT(S): Orbitrap Eclipse, Orbitrap Fusion Lumos

ORGANISM(S): Homo Sapiens (ncbitaxon:9606)

SUBMITTER: Alban Ordureau Rachel Green

PROVIDER: MSV000092521 | MassIVE | Tue Jul 25 07:20:00 BST 2023

REPOSITORIES: MassIVE

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The ribotoxic stress response drives UV-mediated cell death.

Sinha Niladri K NK McKenney Connor C Yeow Zhong Y ZY Li Jeffrey J JJ Nam Ki Hong KH Yaron-Barir Tomer M TM Johnson Jared L JL Huntsman Emily M EM Cantley Lewis C LC Ordureau Alban A Regot Sergi S Green Rachel R

Cell 20240605 14

While ultraviolet (UV) radiation damages DNA, eliciting the DNA damage response (DDR), it also damages RNA, triggering transcriptome-wide ribosomal collisions and eliciting a ribotoxic stress response (RSR). However, the relative contributions, timing, and regulation of these pathways in determining cell fate is unclear. Here we use time-resolved phosphoproteomic, chemical-genetic, single-cell imaging, and biochemical approaches to create a chronological atlas of signaling events activated in ce ...[more]

PMID: 38843833

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Project description:Proteome maps of biochemically enriched synaptic tissue from human Alzheimers Disease (AD) and control angular gyrus samples to highlight the synapse as a site of integration for pathophysiological processes active in AD. The samples were combined into eleven TMTpools (01 to 11). Samples pools were fractionated using basic pH reversed-phase chromatography (bRPLC) and at least 12 fractions were analyzed by LC-MS2/MS3 per pool. The RAW files are: Carlyle_TMTpool_01_fraction_01 to Carlyle_TMTpool_01_fraction_12 Carlyle_TMTpool_02_fraction_01 to Carlyle_TMTpool_02_fraction_12 Carlyle_TMTpool_03_fraction_01 to Carlyle_TMTpool_03_fraction_12 Carlyle_TMTpool_04_fraction_01 to Carlyle_TMTpool_04_fraction_12 Carlyle_TMTpool_05_fraction_01 to Carlyle_TMTpool_05_fraction_12 Carlyle_TMTpool_06_fraction_01 to Carlyle_TMTpool_06_fraction_12 Carlyle_TMTpool_07_fraction_01 to Carlyle_TMTpool_07_fraction_12 Carlyle_TMTpool_08_fraction_01 to Carlyle_TMTpool_08_fraction_08, Carlyle_TMTpool_08_fraction_09A, Carlyle_TMTpool_08_fraction_09B, Carlyle_TMTpool_08_fraction_10 to Carlyle_TMTpool_08_fraction_12 Carlyle_TMTpool_09_fraction_01 to Carlyle_TMTpool_09_fraction_12 Carlyle_TMTpool_10_fraction_01 to Carlyle_TMTpool_10_fraction_12 Carlyle_TMTpool_11_fraction_01 to Carlyle_TMTpool_11_fraction_12 The labeling scheme is: TMTpool_01: RUSH_0592 (126), RUSH_0034 (127n), RUSH_0755 (127c), RUSH_0143 (128n), RUSH_1442 (128c), RUSH_0570 (129n), Pooled_sample (129c), Pooled_sample (130n), RUSH_0992 (130c), RUSH_0857 (131n), bridge (131c). TMTpool_02: RUSH_0974 (126), RUSH_1440 (127n), RUSH_0240 (127c), RUSH_0032 (128n), RUSH_0406 (128c), RUSH_0065 (129n), RUSH_1376 (129c), RUSH_0684 (130n), RUSH_0269 (130c), RUSH_1159 (131n), bridge (131c). TMTpool_03: RUSH_0136 (126), Pooled_sample (127n), RUSH_0537 (127c), RUSH_1250 (128n), RUSH_0795 (128c), RUSH_0001 (129n), RUSH_0404 (129c), RUSH_0606 (130n), RUSH_0229 (130c), RUSH_0465 (131n), bridge (131c). TMTpool_04: RUSH_0734 (126), RUSH_0873 (127n), RUSH_0743 (127c), Pooled_sample (128n), RUSH_0712 (128c), RUSH_0751 (129n), RUSH_0302 (129c), RUSH_0499 (130n), RUSH_1218 (130c), RUSH_1421 (131n), bridge (131c). TMTpool_05: RUSH_1392 (126), RUSH_0890 (127n), RUSH_1006 (127c), RUSH_1063 (128n), RUSH_0128 (128c), RUSH_0590 (129n), RUSH_0674 (129c), RUSH_0572 (130n), RUSH_0413 (130c), RUSH_1288 (131n), bridge (131c). TMTpool_06: RUSH_1487 (126), RUSH_1348 (127n), RUSH_0007 (127c), RUSH_1323 (128n), RUSH_1498 (128c), RUSH_0976 (129n), RUSH_0859 (129c), RUSH_0544 (130n), RUSH_0067 (130c), RUSH_1384 (131n), bridge (131c). TMTpool_07: RUSH_0212 (126), RUSH_1474 (127n), Pooled_sample (127c), RUSH_0010 (128n), RUSH_0931 (128c), RUSH_1205 (129n), RUSH_1200 (129c), RUSH_0206 (130n), RUSH_0376 (130c), RUSH_0460 (131n), bridge (131c). TMTpool_08: RUSH_0927 (126), RUSH_0963 (127n), RUSH_0055 (127c), RUSH_0085 (128n), RUSH_0780 (128c), RUSH_0327 (129n), RUSH_0354 (129c), RUSH_0123 (130n), RUSH_1358 (130c), RUSH_1424 (131n), bridge (131c). TMTpool_09: RUSH_1334 (126), RUSH_0921 (127n), Pooled_sample (127c), RUSH_1093 (128n), RUSH_1499 (128c), RUSH_1419 (129n), Pooled_sample (129c), RUSH_0555 (130n), RUSH_0618 (130c), RUSH_0732 (131n), bridge (131c). TMTpool_10: RUSH_1176 (126), Pooled_sample (127n), RUSH_0958 (127c), RUSH_0447 (128n), RUSH_1485 (128c), RUSH_0158 (129n), RUSH_0252 (129c), RUSH_0078 (130n), RUSH_0415 (130c), Pooled_sample (131n), bridge (131c). TMTpool_11: RUSH_1313 (126), RUSH_0989 (127n), RUSH_0295 (127c), RUSH_0177 (128n), RUSH_0064 (128c), RUSH_0919 (129n), RUSH_0847 (129c), RUSH_0266 (130n), RUSH_0933 (130c), Pooled_sample (131n), bridge (131c).

Project description:We used mass spectrometry-based proteomics to perform a phosphoproteomics analysis of selinexor-treated ex vivo AML patient samples (n=20) and 4 AML cell lines (PL-21, NOMO-1, GDM-1 and MV4-11). For quantitative phosphoproteomics, we used a tandem mass tag (TMT)-based approach. Conditions for the TMT 11-plex (Ex vivo samples) and TMT 8-plex (AML cell lines) setups are specified below. Cells were treated with 1 M selinexor (also known as KPT-330) or 0.1% DMSO for 6 hours. The experimental treatment conditions and TMT11-plex (ex vivo samples) and TMT8-plex (cell lines) labeling schemes are specified below: TMT-11-plex TMT channel Patient no Treatment Set 1 126 Ex_24 DMSO Set 1 127N Ex_24 Selinexor Set 1 127C Ex_25 DMSO Set 1 128N Ex_25 Selinexor Set 1 128C Ex_34 DMSO Set 1 129N Ex_34 Selinexor Set 1 129C Ex_41 DMSO Set 1 130N Ex_41 Selinexor Set 1 130C Ex_44 DMSO Set 1 131N Ex_44 Selinexor Set 1 131C Mix pool Mix pool Set 2 126 Ex_9 DMSO Set 2 127N Ex_9 Selinexor Set 2 127C Ex_18 DMSO Set 2 128N Ex_18 Selinexor Set 2 128C Ex_27 DMSO Set 2 129N Ex_27 Selinexor Set 2 129C Ex_29 DMSO Set 2 130N Ex_29 Selinexor Set 2 130C Ex_36 DMSO Set 2 131N Ex_36 Selinexor Set 2 131C Mix pool Mix pool Set 3 126 Ex_8 DMSO Set 3 127N Ex_8 Selinexor Set 3 127C Ex_13 DMSO Set 3 128N Ex_13 Selinexor Set 3 128C Ex_14 DMSO Set 3 129N Ex_14 Selinexor Set 3 129C Ex_19 DMSO Set 3 130N Ex_19 Selinexor Set 3 130C Ex_40 DMSO Set 3 131N Ex_40 Selinexor Set 3 131C Mix pool Mix pool Set 4 126 Ex_6 DMSO Set 4 127N Ex_6 Selinexor Set 4 127C Ex_15 DMSO Set 4 128N Ex_15 Selinexor Set 4 128C Ex_28 DMSO Set 4 129N Ex_28 Selinexor Set 4 129C Ex_39 DMSO Set 4 130N Ex_39 Selinexor Set 4 130C Ex_42 DMSO Set 4 131N Ex_42 Selinexor Set 4 131C Mix pool Mix pool AML cell lines: TMT-8-plex TMT channel Treatment Replicate GDM-1 126 DMSO 1 GDM-1 127N DMSO 2 GDM-1 127C DMSO 3 GDM-1 128N DMSO 4 GDM-1 128C Selinexor 1 GDM-1 129N Selinexor 2 GDM-1 129C Selinexor 3 GDM-1 130N Selinexor 4 MV4-11 126 DMSO 1 MV4-11 127N DMSO 2 MV4-11 127C DMSO 3 MV4-11 128N DMSO 4 MV4-11 128C Selinexor 1 MV4-11 129N Selinexor 2 MV4-11 129C Selinexor 3 MV4-11 130N Selinexor 4 NOMO-1 126 DMSO 1 NOMO-1 127N DMSO 2 NOMO-1 127C DMSO 3 NOMO-1 128N DMSO 4 NOMO-1 128C Selinexor 1 NOMO-1 129N Selinexor 2 NOMO-1 129C Selinexor 3 NOMO-1 130N Selinexor 4 PL-21 126 DMSO 1 PL-21 127N DMSO 2 PL-21 127C DMSO 3 PL-21 128N DMSO 4 PL-21 128C Selinexor 1 PL-21 129N Selinexor 2 PL-21 129C Selinexor 3 PL-21 130N Selinexor 4

Project description:Three flasks of asynchronous blood stage P. falciparum Dd2 culture were grown to ~15% parasitemia, 4% hematocrit using the Trager- Jensen method. Per biological replicate, parasites were treated with 5 x EC50 or EC90 of compound depending on the slope of the EC50 curve for 3 hours. Compounds are DC-12717, DC-12834, BI-2536, GSK461364, and DC-12862. All compounds have three biological replicates, except DC-12862 which has one. As a control, culture was treated with an equal volume of DMSO vehicle. Parasites were collected through saponin lysis of erythrocytes, and parasite pellets were resuspended in 8 M urea lysis buffer (8 M urea, 50 mM Tris pH 8.5, 1% SDS), supplemented with a protease and phosphatase inhibitor cocktail. Parasites were lysed by sonication, and protein was quantified with the BCA protein assay. Samples were reduced, alkylated, resuspended, and digested. Resulting peptides were quantified and labelled with isobaric tandem mass tags (TMT). Following a ratio check, phospho-enrichment was performed with a Pierce High-Select Fe-NTA Phosphopeptide Enrichment Kit. The resultant phosphopeptide eluate was analyzed with LC-MS2. The peptide flowthrough was fractionated by basic reverse phase HPLC and similarly analyzed with LC-MS3. Spectra were searched against a composite P. falciparum and human Uniprot database (https://www.uniprot.org/), using the COMET algorithm. Peptide matches were filtered with a 1% false discovery rate (FDR), and peptides with a summed signal-to-noise (SN) threshold of >100 were quantified. For phosphopeptides, a 1% FDR filter was used, and a summed SN threshold of greater than or equal to 100. Channels: 126 - Culture 1 Control, 127N - Culture 2 Control, 127C - Culture 3 Control, 128N - Culture 1 DC-12717 128C - Culture 2 DC-12717 129N - Culture 3 DC-12717, 129C - Culture 1 DC-12835, 130N - Culture 2 DC-12835, 130C - Culture 3 DC-12835, 131N - Culture 1 BI-2536, 131C - Culture 2 BI-2536, 132N - Culture 3 BI-2536, 132C - Culture 1 GSK461364, 133N - Culture 2 GSK461364, 133C - Culture 3 GSK461364, 134N - Culture 1 DC-12862

Project description:Mass spectrometry-based proteomics was applied to unravel Erk signaling in mouse embryonic stem cells (ESCs). A previously described engineered ESCs system for Erk induction via a drug-inducible cRAF-ErT2 fusion (Hamilton, W. B. & Brickman, J. M., Cell Rep 2014) was used in two different setups measuring the phosphoproteome in an 11-plex tandem mass tag (TMT)-based approach as described below. Fgf4 stimulation and MEK inhibition: ESCs were stimulated with Fgf4 (40 nM, recombinant, mouse, 5 minutes) following pretreatment (30 minutes) with either DMSO (1:10000) or MEK inhibition (MEKi: 1 microM PD0325901): The experimental treatment conditions and TMT11-plex labeling are specified below: 126: DMSO, unstimulated control replicate 1 127N: DMSO, unstimulated control replicate 2 127C: DMSO, unstimulated control replicate 3 128N: DMSO + Fgf4 replicate 1 128C: DMSO + Fgf4 replicate 2 129N: DMSO + Fgf4 replicate 3 129C: MEKi + Fgf4 replicate 1 130N: MEKi + Fgf4 replicate 2 130C: MEKi + Fgf4 replicate 3 131N: MEKi replicate 1 131C: MEKi replicate 2 Analog sensitive Erk2 setup: In the ESC system the ERK2-/- ESCs (Hamilton et al., PloS ONE, 2013) were engineered to stably express an inducible cRAF-ErT2 fusion protein expressed from the same transgene as a FLAG-tagged analogue sensitive Erk2Q103A. Erk1 was subsequently removed by CRISPR-Cas9 mutagenesis using sgRNAs flanking exon 2. Cells were pre-treated for 30 minutes with either the ATP-mimetic compound 1-NM-PP1 (2 microM) (Endo et al., Biochem Biophys Res Commun 2006), the Mek1 inhibitor PD0325901 (1 microM), or DMSO (0.01%), followed by induction or not of the cRAF-ErT2 fusion by (Z)-4-Hydroxytamoxifen (4OHT) (250 nM) for 2 hours as indicated. The experimental treatment conditions and TMT11-plex labeling are specified below: 126: DMSO pretreatment, non-induced control, replicate 1 127N: DMSO pretreatment, non-induced control, replicate 2 127C: DMSO pretreatment, non-induced control, replicate 3 128N: DMSO pretreatment, 4OHT-induction, replicate 1 128C: DMSO pretreatment, 4OHT-induction, replicate 2 129N: DMSO pretreatment, 4OHT-induction, replicate 3 129C: PP1 pretreatment, 4OHT-induction, replicate 1 130N: PP1 pretreatment, 4OHT-induction, replicate 2 130C: PP1 pretreatment, 4OHT-induction, replicate 3 131N: MEKi pretreatment, 4OHT-induction, replicate 1 131C: MEKi pretreatment, 4OHT-induction, replicate 2

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The ribotoxic stress response drives UV-mediated cell death.

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The ribotoxic stress response drives UV-mediated cell death.

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