title: MERTK arginine-844-cysteine in a patient with severe rod-cone dystrophy: loss of mutant protein function in transfected cells.
abstract: OBJECTIVE
  Mutations in the MERTK gene are responsible for retinal degeneration in the Royal College of Surgeons (RCS) rat and are a cause of human autosomal recessive retinitis pigmentosa (RP). This study reports the identification and functional analysis of novel MERTK mutations to provide information regarding whether they are causative of severe rod-cone degeneration in a young patient.
  
  
  METHODS
  MERTK missense variants identified by single-strand conformational polymorphism (SSCP) and sequence analysis were introduced into expression constructs and used to transfect HEK293T cells. Recombinant protein expression was assayed with anti-MERTK and anti-phosphotyrosine antibodies. Protein turnover was assayed in pulse-chase studies of 35S-methionine incorporation. Transcript levels were determined by quantitative RT-PCR.
  
  
  RESULTS
  Three MERTK sequence variants were identified in a patient with rod-cone dystrophy: R722X in exon 16 and R865W in exon 19 on the paternal allele and R844C in exon 19 on the maternal allele. The R844C sequence change affects an evolutionarily conserved amino acid residue and was not detected in unaffected individuals. In transfected HEK293Tcells, wild-type (wt) and W865 MERTK were expressed at equivalent levels and present in the plasma membrane, stimulated tyrosine phosphorylation, and induced significant rounding of the cell bodies. In contrast, C844 MERTK was expressed at low levels and did not stimulate tyrosine phosphorylation. In addition, the relative stability of C844 MERTK was significantly less than wt in assays of protein turnover. At age 13, the patient had 20/60 and 20/200 acuities, tunnel vision of 5 degrees centrally, and a far temporal peripheral crescent bilaterally, and ERGs were nondetectable. The fundi showed bull's-eye macular atrophy and widespread RPE thinning.
  
  
  CONCLUSIONS
  The present study reports the identification of R844C, the first putative pathogenic MERTK missense mutation that results in severe retinal degeneration with childhood onset when in compound heterozygous form with a R722X allele. The loss of function of C844 MERTK is probably due to decreased protein stability.
authors.affiliation: Department of Ophthalmology and Visual Sciences, University of Michigan Medical School, Ann Arbor, Michigan 48105, USA.
authors.forename: Christina L
authors.initials: CL
authors.lastname: McHenry
authors.forename: Yuhui
authors.initials: Y
authors.lastname: Liu
authors.forename: Wei
authors.initials: W
authors.lastname: Feng
authors.forename: Anita R
authors.initials: AR
authors.lastname: Nair
authors.forename: Kecia L
authors.initials: KL
authors.lastname: Feathers
authors.forename: Xiaoling
authors.initials: X
authors.lastname: Ding
authors.forename: Andreas
authors.initials: A
authors.lastname: Gal
authors.forename: Douglas
authors.initials: D
authors.lastname: Vollrath
authors.forename: Paul A
authors.initials: PA
authors.lastname: Sieving
authors.forename: Debra A
authors.initials: DA
authors.lastname: Thompson
mesh_terms: D000293:Adolescent; D001120:Arginine; D015153:Blotting, Western; D003545:Cysteine; D005260:Female; D005786:Gene Expression Regulation / Q000502:physiology*; D014644:Genetic Variation; D006801:Humans; D007668:Kidney / Q000196:embryology / Q000378:metabolism; D020125:Mutation, Missense; D010766:Phosphorylation; D020419:Photoreceptor Cells, Vertebrate / Q000473:pathology*; D018807:Polymorphism, Single-Stranded Conformational; D011518:Proto-Oncogene Proteins; D020794:Receptor Protein-Tyrosine Kinases / Q000235:genetics* / Q000378:metabolism; D012162:Retinal Degeneration / Q000235:genetics* / Q000378:metabolism / Q000473:pathology; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014162:Transfection; D014443:Tyrosine / Q000378:metabolism; D000076202:c-Mer Tyrosine Kinase
chemical_list: D011518:Proto-Oncogene Proteins; D014443:Tyrosine; D001120:Arginine; C097072:MERTK protein, human; C490765:Mertk protein, rat; D020794:Receptor Protein-Tyrosine Kinases; D000076202:c-Mer Tyrosine Kinase; D003545:Cysteine