title: Alternative splicing in osteoclasts and Paget's disease of bone.
abstract: BACKGROUND
  Mutations in the SQSTM1/p62 gene have been reported in Paget's disease of bone (PDB), but they are not sufficient to induce the pagetic osteoclast (OC) phenotype. We hypothesized that specific RNA isoforms of OC-related genes may contribute to the overactivity of pagetic OCs, along with other genetic predisposing factors.
  
  
  METHODS
  Alternative splicing (AS) events were studied using a PCR-based screening strategy in OC cultures from 29 patients with PDB and 26 healthy donors (HD), all genotyped for the p62P392L mutation. Primer pairs targeting 5223 characterized AS events were used to analyze relative isoform ratios on pooled cDNA from samples of the four groups (PDB, PDBP392L, HD, HDP392L). Of the 1056 active AS events detected in the screening analysis, 192 were re-analyzed on non-amplified cDNA from each subject of the whole cohort.
  
  
  RESULTS
  This analysis led to the identification of six AS events significantly associated with PDB, but none with p62P392L. The corresponding genes included LGALS8, RHOT1, CASC4, USP4, TBC1D25, and PIDD. In addition, RHOT1 and LGALS8 genes were upregulated in pagetic OCs, as were CASC4 and RHOT1 genes in the presence of p62P392L. Finally, we showed that the proteins encoded by LGALS8, RHOT1, USP4, TBC1D25, and PIDD were expressed in human OCs.
  
  
  CONCLUSIONS
  This study allowed the identification of hitherto unknown players in OC biology, and our findings of a differential AS in pagetic OCs may generate new concepts in the pathogenesis of PDB.
authors.forename: Roscoe
authors.initials: R
authors.lastname: Klinck
authors.forename: Gino
authors.initials: G
authors.lastname: Laberge
authors.forename: Martine
authors.initials: M
authors.lastname: Bisson
authors.forename: Stephen
authors.initials: S
authors.lastname: McManus
authors.forename: Laëtitia
authors.initials: L
authors.lastname: Michou
authors.forename: Jacques P
authors.initials: JP
authors.lastname: Brown
authors.forename: Sophie
authors.initials: S
authors.lastname: Roux
mesh_terms: D048868:Adaptor Proteins, Signal Transducing / Q000235:genetics; D017398:Alternative Splicing; D002478:Cells, Cultured; D005260:Female; D020869:Gene Expression Profiling; D005786:Gene Expression Regulation; D005838:Genotype; D006801:Humans; D008297:Male; D009154:Mutation; D010001:Osteitis Deformans / Q000235:genetics* / Q000473:pathology; D010010:Osteoclasts / Q000473:pathology*; D012313:RNA / Q000032:analysis; D000071456:Sequestosome-1 Protein
chemical_list: D048868:Adaptor Proteins, Signal Transducing; C099718:SQSTM1 protein, human; D000071456:Sequestosome-1 Protein; D012313:RNA