PXD001485

PXD001485 is an original dataset announced via ProteomeXchange.

Title	FOXP1 is a transcriptional Coactivator of Wnt/β-catenin signaling in Diffuse Large B-cell Lymphoma
Description	We used a mass spectrometry-coupled lentiviral ‘CD-tagging’ mutagenesis approach to identify genes that activate Wnt/β-catenin signaling. Human A375 melanoma cells containing a β-catenin-driven GFP (green fluorescent protein) transcriptional reporter were transduced with CDBF lentivirus. When integrated near an expressed and spliced gene, the cytomegalovirus (CMV) promoter of the CDBF vector drives constitutive BFP (blue fluorescence protein) expression and by virtue of the splice donor (SD) sequence, an overexpressed FLAG-tagged fusion of the targeted gene. Depending on where within the gene locus the CDBP vector integrates, the resulting overexpressed gene product may be full length or amino-terminal trunctated. Fluorescence activated cell sorting (FACS) was used to isolate BFP+/GFP+ (Wnt active) or BFP+/GFP- (Wnt inactive) A375 cells. We reasoned that if successful, FLAG epitope tag immunopurification and mass spectrometry-based identification of the overexpressed fusion proteins would be cheaper, faster and provide more information than traditional PCR-based detection. Five FLAG immunopurification followed by a series of high salt washes, on-bead tryptic digestion and shotgun mass spectrometry (MS) identified 20 high-confidence proteins specific to Wnt-active cells. The high-salt washes removed associated proteins from the FLAG-tagged bait proteins. The FOXP1 transcription factor ranked as the top screen hit, as determined by spectral count abundance and the CompPASS WD-score.
HostingRepository	PRIDE
AnnounceDate	2016-07-05
AnnouncementXML	Submission_2016-07-05_05:58:48.xml
DigitalObjectIdentifier
ReviewLevel	Peer-reviewed dataset
DatasetOrigin	Original dataset
RepositorySupport	Unsupported dataset by repository
PrimarySubmitter	Dennis Goldfarb
SpeciesList	scientific name: Homo sapiens (Human); NCBI TaxID: 9606;
ModificationList	monohydroxylated residue; iodoacetamide derivatized residue
Instrument	LTQ Orbitrap Velos

Revision	Datetime	Status	ChangeLog Entry
0	2014-11-13 01:02:10	ID requested
⏵ 1	2016-07-05 05:58:49	announced

Walker MP, Stopford CM, Cederlund M, Fang F, Jahn C, Rabinowitz AD, Goldfarb D, Graham DM, Yan F, Deal AM, Fedoriw Y, Richards KL, Davis IJ, Weidinger G, Damania B, Major MB, -catenin signaling in diffuse large B cell lymphoma. Sci Signal, 8(362):ra12(2015) [pubmed]

curator keyword: Biomedical

submitter keyword: FOXP1, Gain-of-function screening, FLAG APMS, Wnt signaling

Michael Ben Major
contact affiliation	Department of Cell Biology and Physiology, Lineberger Comprehensive Cancer Center, University of North Carolina at Chapel Hill, Chapel Hill, North Carolina, USA.
contact email	ben_major@med.unc.edu
lab head
Dennis Goldfarb
contact affiliation	Cell and Developmental Biology
contact email	dennisg@email.unc.edu
dataset submitter

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PRIDE project URI

• PRIDE
  1. PXD001485
     1. Label: PRIDE project
     2. Name: FOXP1 is a transcriptional Coactivator of Wnt/β-catenin signaling in Diffuse Large B-cell Lymphoma