title: Transcript mutations of the alpha regulatory subunit of protein kinase A and up-regulation of the RNA-editing gene transcript in lupus T lymphocytes.
abstract: BACKGROUND
  Systemic lupus erythematosus (SLE) is an autoimmune disorder characterised by diverse dysfunctions of immune effector cells, including proliferation and cytotoxicity. In T cells from patients with SLE, activity of type 1 protein kinase A isozymes is greatly reduced because of decreased expression of the alpha and beta regulatory subunits (RI alpha and RI beta). We aimed to identify a molecular mechanism or mechanisms for this isozyme deficiency by assessing occurrence of mutations in transcripts of the RI alpha subunit in patients with SLE.
  
  
  METHODS
  We cloned and sequenced cDNA of RI alpha and corresponding genomic DNA of the coding region to detect sequence changes from eight patients with SLE and six healthy controls. Because transcript editing is regulated by adenosine deaminases that act on RNA (ADAR), we quantified expression of ADAR1 transcripts in SLE and control T cells by competitive PCR.
  
  
  RESULTS
  Sequence analyses of cDNA showed heterogeneous transcript mutations, including deletions, transitions, and transversions. We identified 1.22 x 10(-3)/bp transcript mutations in SLE T cells-a frequency 7.5 times higher than that in control T cells. By contrast, we identified no genomic mutations. Two hotspots were identified in the RI alpha subunit transcripts from SLE T cells, one located adjacent to a pseudosubstrate site of the RI alpha subunit and the other a component of the cAMP binding A domain. ADAR1 mRNA content was 3.5 times higher in SLE cells than in control T cells (p=0.001).
  
  
  CONCLUSIONS
  An RNA-editing enzyme could be converting adenosine to inosine within double-stranded regions of RNA, resulting in transcript mutations. This process could be one mechanism resulting in mutations in the RI alpha subunit of type 1 protein kinase A.
authors.affiliation: Section on Rheumatology and Clinical Immunology, Department of Internal Medicine, Wake Forest University School of Medicine, Winston-Salem, NC 27157, USA. dlaxmina@wfubmc.edu
authors.forename: Dama
authors.initials: D
authors.lastname: Laxminarayana
authors.forename: Islam U
authors.initials: IU
authors.lastname: Khan
authors.forename: Gary
authors.initials: G
authors.lastname: Kammer
mesh_terms: D000243:Adenosine Deaminase / Q000235:genetics; D000328:Adult; D001483:Base Sequence / Q000235:genetics; D015331:Cohort Studies; D054756:Cyclic AMP-Dependent Protein Kinase RIalpha Subunit; D017868:Cyclic AMP-Dependent Protein Kinases / Q000235:genetics*; D004252:DNA Mutational Analysis; D018076:DNA, Complementary / Q000235:genetics; D005260:Female; D015870:Gene Expression / Q000502:physiology; D005787:Gene Frequency; D006801:Humans; D008180:Lupus Erythematosus, Systemic / Q000235:genetics* / Q000276:immunology; D008297:Male; D009154:Mutation / Q000235:genetics*; D017393:RNA Editing / Q000235:genetics*; D012333:RNA, Messenger / Q000235:genetics; D016601:RNA-Binding Proteins; D034743:RNA-Induced Silencing Complex; D012261:Ribonucleoproteins; D013601:T-Lymphocytes / Q000378:metabolism*; D014158:Transcription, Genetic / Q000235:genetics*; D015854:Up-Regulation / Q000235:genetics
chemical_list: D054756:Cyclic AMP-Dependent Protein Kinase RIalpha Subunit; D018076:DNA, Complementary; C517834:PRKAR1A protein, human; D012333:RNA, Messenger; D016601:RNA-Binding Proteins; D034743:RNA-Induced Silencing Complex; D012261:Ribonucleoproteins; D017868:Cyclic AMP-Dependent Protein Kinases; C567955:ADARB1 protein, human; D000243:Adenosine Deaminase