title: Morphological analysis of 13 LMNA variants identified in a cohort of 324 unrelated patients with idiopathic or familial dilated cardiomyopathy.
abstract: BACKGROUND
  Mutations in the LMNA gene, encoding lamins A/C, represent a significant cause of dilated cardiomyopathy. We recently identified 18 protein-altering LMNA variants in a cohort of 324 unrelated patients with dilated cardiomyopathy. However, at least one family member with dilated cardiomyopathy in each of 6 pedigrees lacked the LMNA mutation (nonsegregation), whereas small sizes of 5 additional families precluded definitive determinations of segregation, raising questions regarding contributions by those variants to disease.
  
  
  RESULTS
  We have consequently expressed, in COS7 cells, GFP-prelamin A (GFPLaA) fusion constructs incorporating the 6 variants in pedigrees with nonsegregation (R101P, A318T, R388H, R399C, S437Hfsx1, and R654X), the 4 variants in pedigrees with unknown segregation (R89L, R166P [in 2 families], I210S, R471H), and 3 additional missense variants (R190Q, E203K, and L215P) that segregated with disease. Confocal immunofluorescence microscopy was used to characterize GFP-lamin A localization and nuclear morphology. Abnormal phenotypes were observed for 10 of 13 (77%) variants (R89L, R101P, R166P, R190Q, E203K, I210S, L215P, R388H, S437Hfsx1, and R654X), including 4 of 6 showing nonsegregation and 3 of 4 with uncertain segregation. All 7 variants affecting coil 1B and the lamin A-only mutation, R654X, exhibited membrane-bound GFP-lamin A aggregates and nuclear shape abnormalities. Unexpectedly, R388H largely restricted GFP-lamin A to the cytoplasm. Equally unexpected were unique streaked aggregates with S437Hfsx1 and giant aggregates with both S437Hfsx1 and R654X.
  
  
  CONCLUSIONS
  This work expands the recognized spectrum of lamin A localization abnormalities in dilated cardiomyopathy. It also provides evidence supporting pathogenicity of 10 of 13 tested LMNA variants, including some with uncertain or nonsegregation.
authors.affiliation: Cardiovascular Division, Department of Medicine, University of Miami Miller School of Medicine, Miami, FL 33136, USA.
authors.forename: Jason
authors.initials: J
authors.lastname: Cowan
authors.forename: Duanxiang
authors.initials: D
authors.lastname: Li
authors.forename: Jorge
authors.initials: J
authors.lastname: Gonzalez-Quintana
authors.forename: Ana
authors.initials: A
authors.lastname: Morales
authors.forename: Ray E
authors.initials: RE
authors.lastname: Hershberger
mesh_terms: D019943:Amino Acid Substitution; D000818:Animals; D019556:COS Cells; D002311:Cardiomyopathy, Dilated / Q000235:genetics* / Q000473:pathology; D002467:Cell Nucleus / Q000648:ultrastructure; D002522:Chlorocebus aethiops; D015331:Cohort Studies; D005260:Female; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D006801:Humans; D034904:Lamin Type A / Q000235:genetics* / Q000378:metabolism; D008297:Male; D016297:Mutagenesis, Site-Directed; D009687:Nuclear Proteins / Q000235:genetics; D010375:Pedigree; D010641:Phenotype; D011498:Protein Precursors / Q000235:genetics; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism
chemical_list: D034904:Lamin Type A; D009687:Nuclear Proteins; D011498:Protein Precursors; D011993:Recombinant Fusion Proteins; C090764:prelamin A; D049452:Green Fluorescent Proteins