title: BMPR1A mutations in juvenile polyposis affect cellular localization.
abstract: BACKGROUND
  Juvenile polyposis (JP) is characterized by the development of hamartomatous polyps of the gastrointestinal tract that collectively carry a significant risk of malignant transformation. Mutations in the bone morphogenetic protein receptor type 1A (BMPR1A) are known to predispose to JP. We set out to study the effect of such missense mutations on BMPR1A cellular localization.
  
  
  METHODS
  We chose eight distinct mutations for analysis. We tagged a BMPR1A wild-type (WT) expression plasmid with green fluorescent protein on its C-terminus. Site-directed mutagenesis was used to recreate JP patient mutations from the WT-green fluorescent protein BMPR1A plasmid. We verified mutant expression vector sequences by direct sequencing. First, we transfected BMPR1A expression vectors into HEK-293T cells; then, we performed confocal microscopy to determine cellular localization. Four independent observers used a scoring system from 1 to 3 to categorize the degree of membrane versus cellular localization.
  
  
  RESULTS
  Of the eight selected mutations, one was within the signaling peptide, four were within the extracellular domain, and three were within the intracellular domain. The WT BMPR1A vector had strong membrane staining, whereas all eight mutations had much less membrane and much more intracellular localization. Enzyme-linked immunosorbent assays for BMPR1A demonstrated no significant differences in protein quantities between constructs, except for one affecting the start codon.
  
  
  CONCLUSIONS
  Bone morphogenetic protein receptor type 1A missense mutations occurring in patients with JP affected cellular localization in an in vitro model. These findings suggest a mechanism by which such mutations can lead to disease by altering downstream signaling through the bone morphogenetic protein pathway.
authors: Division of Surgical Oncology and Endocrine Surgery, Department of Surgery, University of Iowa Hospitals and Clinics, Iowa City, Iowa. Electronic address: james-howe@uiowa.edu.
authors: James R
authors: JR
authors: Howe
authors: Fadi S
authors: FS
authors: Dahdaleh
authors: Jennifer C
authors: JC
authors: Carr
authors: Donghong
authors: D
authors: Wang
authors: Scott K
authors: SK
authors: Sherman
authors: James R
authors: JR
authors: Howe
mesh_terms: D052005:Bone Morphogenetic Protein Receptors, Type I / Q000235:genetics* / Q000378:metabolism*; D002462:Cell Membrane / Q000378:metabolism* / Q000473:pathology; D049452:Green Fluorescent Proteins; D057809:HEK293 Cells; D006801:Humans; D044483:Intestinal Polyposis / Q000151:congenital* / Q000235:genetics* / Q000378:metabolism; D042541:Intracellular Space / Q000378:metabolism*; D007668:Kidney / Q000378:metabolism / Q000473:pathology; D018613:Microscopy, Confocal; D020125:Mutation, Missense / Q000235:genetics*; D009386:Neoplastic Syndromes, Hereditary / Q000235:genetics* / Q000378:metabolism / Q000473:pathology; D015398:Signal Transduction / Q000502:physiology; D014162:Transfection
chemical_list: D049452:Green Fluorescent Proteins; C498211:BMPR1A protein, human; D052005:Bone Morphogenetic Protein Receptors, Type I
keywords: Bone morphogenetic protein; Juvenile polyposis; Missense