Dataset Information

Human cell lines pulldown interactions CL-MSMS

ABSTRACT: Using a combination of GFP-trap affinity purification, native gel band identification and APEX2-mediated BioID pulse-chase analysis, we investigate an import pathway and folding mechanism for histones H3 and H4. We investigated proteins associating with H3 and H4 bearing mutations along alpha-helix 2 using affinity-capture followed by on-beads trypsin digestion and label-free mass spectrometry. In addition, we investigated the composition subcomplexes of affinity-purified histone chaperone NASP separated by native-PAGE, followed by in-gel trypsin digestion and label-free mass spectrometry. Finally, we investigated the proximity in vivo to histone H3 construct under a pulse-chase system (RAPID-release) using two different time points for APEX2-based biotin-phenol biotinylation, 0 minutes and 120 minutes. Samples were quenched and streptavidin-purified, followed by on-beads digestion and label-free mass spectrometry.

INSTRUMENT(S): timsTOF Pro, Orbitrap Fusion

ORGANISM(S): Homo Sapiens (human)

SUBMITTER: Andrew Bottrill

LAB HEAD: Andrew Bowman

PROVIDER: PXD029354 | Pride | 2022-09-15

REPOSITORIES: Pride

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Dataset's files

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			Action	DRS
	Experiment_MS19-054_Alonso_1104_1_from_OT_190319_102_MS19-054_1104_1.mzid.gz	Mzid
	Experiment_MS19-054_Alonso_1104_1_from_OT_190319_102_MS19-054_1104_1.mzid_Experiment_MS19-054_Alonso_1104_1_from_OT_190319_102_MS19-054_1104_1.MGF	Mzid
	Experiment_MS19-054_Alonso_1104_2_from_OT_190319_102_MS19-054_1104_2.mzid.gz	Mzid
	Experiment_MS19-054_Alonso_1104_2_from_OT_190319_102_MS19-054_1104_2.mzid_Experiment_MS19-054_Alonso_1104_2_from_OT_190319_102_MS19-054_1104_2.MGF	Mzid
	Experiment_MS19-054_Alonso_1104_3_from_OT_190319_102_MS19-054_1104_3.mzid.gz	Mzid

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Publications

A specific role for importin-5 and NASP in the import and nuclear hand-off of monomeric H3.

Pardal Alonso Javier AJ Bowman Andrew James AJ

eLife 20220906

Core histones package chromosomal DNA and regulate genomic transactions, with their nuclear import and deposition involving importin-β proteins and a dedicated repertoire of histone chaperones. Previously, a histone H3-H4 dimer has been isolated bound to importin-4 (Imp4) and the chaperone ASF1, suggesting that H3 and H4 fold together in the cytoplasm before nuclear import. However, other studies have shown the existence of monomeric H3 in the nucleus, indicating a post-import folding pathway. H ...[more]

PMID: 36066346

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