title: N-terminally truncated FOXP1 protein expression and alternate internal FOXP1 promoter usage in normal and malignant B cells.
abstract: Strong FOXP1 protein expression is a poor risk factor in diffuse large B-cell lymphoma and has been linked to an activated B-cell-like subtype, which preferentially expresses short FOXP1 (FOXP1S) proteins. However, both short isoform generation and function are incompletely understood. Here we prove by mass spectrometry and N-terminal antibody staining that FOXP1S proteins in activated B-cell-like diffuse large B-cell lymphoma are N-terminally truncated. Furthermore, a rare strongly FOXP1-expressing population of normal germinal center B cells lacking the N-terminus of the regular long protein (FOXP1L) was identified. Exon-targeted silencing and transcript analyses identified three alternate 5' non-coding exons [FOXP1-Ex6b(s), FOXP1-Ex7b and FOXP1-Ex7c], downstream of at least two predicted promoters, giving rise to FOXP1S proteins. These were differentially controlled by B-cell activation and methylation, conserved in murine lymphoma cells, and significantly correlated with FOXP1S protein expression in primary diffuse large B-cell lymphoma samples. Alternatively spliced isoforms lacking exon 9 (e.g. isoform 3) did not encode FOXP1S, and an alternate long human FOXP1 protein (FOXP1AL) likely generated from a FOXP1-Ex6b(L) transcript was detected. The ratio of FOXP1L:FOXP1S isoforms correlated with differential expression of plasmacytic differentiation markers in U-2932 subpopulations, and altering this ratio was sufficient to modulate CD19 expression in diffuse large B-cell lymphoma cell lines. Thus, the activity of multiple alternate FOXP1 promoters to produce multiple protein isoforms is likely to regulate B-cell maturation.
authors.affiliation: Nuffield Division of Clinical Laboratory Sciences, Radcliffe Department of Medicine, Oxford University, London, UK.
authors.forename: Philip J
authors.initials: PJ
authors.lastname: Brown
authors.affiliation: Nuffield Division of Clinical Laboratory Sciences, Radcliffe Department of Medicine, Oxford University, London, UK.
authors.forename: Duncan M
authors.initials: DM
authors.lastname: Gascoyne
authors.affiliation: Nuffield Division of Clinical Laboratory Sciences, Radcliffe Department of Medicine, Oxford University, London, UK.
authors.forename: Linden
authors.initials: L
authors.lastname: Lyne
authors.affiliation: Nuffield Division of Clinical Laboratory Sciences, Radcliffe Department of Medicine, Oxford University, London, UK.
authors.forename: Hayley
authors.initials: H
authors.lastname: Spearman
authors.affiliation: Nuffield Division of Clinical Laboratory Sciences, Radcliffe Department of Medicine, Oxford University, London, UK.
authors.forename: Suet Ling
authors.initials: SL
authors.lastname: Felce
authors.affiliation: Immunity and Cancer Laboratory, The Francis Crick Institute, Lincoln's Inn Fields Laboratory, Lincoln's Inn Fields, London, UK.
authors.forename: Nora
authors.initials: N
authors.lastname: McFadden
authors.affiliation: Computational Biology Laboratory, The Francis Crick Institute, Lincoln's Inn Fields Laboratory, Lincoln's Inn Fields, London, UK.
authors.forename: Probir
authors.initials: P
authors.lastname: Chakravarty
authors.affiliation: Leeds Teaching Hospitals NHS Trust, HMDS, Leeds Cancer Centre, Kings College London, UK.
authors.forename: Sharon
authors.initials: S
authors.lastname: Barrans
authors.affiliation: Centre of Excellence for Mass Spectrometry, Institute of Psychiatry, Psychology and Neuroscience, Kings College London, UK.
authors.forename: Steven
authors.initials: S
authors.lastname: Lynham
authors.affiliation: Immunity and Cancer Laboratory, The Francis Crick Institute, Lincoln's Inn Fields Laboratory, Lincoln's Inn Fields, London, UK Peter Gorer Department of Immunobiology, Kings College London, UK.
authors.forename: Dinis P
authors.initials: DP
authors.lastname: Calado
authors.affiliation: Centre of Excellence for Mass Spectrometry, Institute of Psychiatry, Psychology and Neuroscience, Kings College London, UK.
authors.forename: Malcolm
authors.initials: M
authors.lastname: Ward
authors.affiliation: Nuffield Division of Clinical Laboratory Sciences, Radcliffe Department of Medicine, Oxford University, London, UK alison.banham@ndcls.ox.ac.uk.
authors.forename: Alison H
authors.initials: AH
authors.lastname: Banham
mesh_terms: D017398:Alternative Splicing; D000818:Animals; D018941:Antigens, CD19 / Q000235:genetics / Q000378:metabolism; D001402:B-Lymphocytes / Q000378:metabolism*; D045744:Cell Line, Tumor; D005091:Exons; D051858:Forkhead Transcription Factors / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D015972:Gene Expression Regulation, Neoplastic; D006801:Humans; D008213:Lymphocyte Activation / Q000235:genetics / Q000276:immunology; D016403:Lymphoma, Large B-Cell, Diffuse / Q000235:genetics* / Q000473:pathology; D051379:Mice; D011401:Promoter Regions, Genetic; D054730:Protein Interaction Domains and Motifs / Q000235:genetics*; D020033:Protein Isoforms; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D012097:Repressor Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*
chemical_list: D018941:Antigens, CD19; C486626:FOXP1 protein, human; D051858:Forkhead Transcription Factors; D020033:Protein Isoforms; D012333:RNA, Messenger; D012097:Repressor Proteins