title: A novel mutation in DDR2 causing spondylo-meta-epiphyseal dysplasia with short limbs and abnormal calcifications (SMED-SL) results in defective intra-cellular trafficking.
abstract: BACKGROUND
  The rare autosomal genetic disorder, Spondylo-meta-epiphyseal dysplasia with short limbs and abnormal calcifications (SMED-SL), is reported to be caused by missense or splice site mutations in the human discoidin domain receptor 2 (DDR2) gene. Previously our group has established that trafficking defects and loss of ligand binding are the underlying cellular mechanisms of several SMED-SL causing mutations. Here we report the clinical characteristics of two siblings of consanguineous marriage with suspected SMED-SL and identification of a novel disease-causing mutation in the DDR2 gene.
  
  
  METHODS
  Clinical evaluation and radiography were performed to evaluate the patients. All the coding exons and splice sites of the DDR2 gene were sequenced by Sanger sequencing. Subcellular localization of the mutated DDR2 protein was determined by confocal microscopy, deglycosylation assay and Western blotting. DDR2 activity was measured by collagen activation and Western analysis.
  
  
  RESULTS
  In addition to the typical features of SMED-SL, one of the patients has an eye phenotype including visual impairment due to optic atrophy. DNA sequencing revealed a novel homozygous dinucleotide deletion mutation (c.2468_2469delCT) on exon 18 of the DDR2 gene in both patients. The mutation resulted in a frameshift leading to an amino acid change at position S823 and a predicted premature termination of translation (p.S823Cfs*2). Subcellular localization of the mutant protein was analyzed in mammalian cell lines, and it was found to be largely retained in the endoplasmic reticulum (ER), which was further supported by its N-glycosylation profile. In keeping with its cellular mis-localization, the mutant protein was found to be deficient in collagen-induced receptor activation, suggesting protein trafficking defects as the major cellular mechanism underlying the loss of DDR2 function in our patients.
  
  
  CONCLUSIONS
  Our results indicate that the novel mutation results in defective trafficking of the DDR2 protein leading to loss of function and disease. This confirms our previous findings that DDR2 missense mutations occurring at the kinase domain result in retention of the mutant protein in the ER.
authors.forename: Adila
authors.initials: A
authors.lastname: Al-Kindi
authors.forename: Praseetha
authors.initials: P
authors.lastname: Kizhakkedath
authors.forename: Huifang
authors.initials: H
authors.lastname: Xu
authors.forename: Anne
authors.initials: A
authors.lastname: John
authors.forename: Abeer Al
authors.initials: AA
authors.lastname: Sayegh
authors.forename: Anuradha
authors.initials: A
authors.lastname: Ganesh
authors.forename: Maha
authors.initials: M
authors.lastname: Al-Awadi
authors.forename: Lamya
authors.initials: L
authors.lastname: Al-Anbouri
authors.forename: Lihadh
authors.initials: L
authors.lastname: Al-Gazali
authors.forename: Birgit
authors.initials: B
authors.lastname: Leitinger
authors.affiliation: Department of Pathology, College of Medicine and Heath Sciences, United Arab Emirates University, P,O, Box 17666 Al Ain, United Arab Emirates. bassam.ali@uaeu.ac.ae.
authors.forename: Bassam R
authors.initials: BR
authors.lastname: Ali
mesh_terms: D001483:Base Sequence; D015153:Blotting, Western; D017931:DNA Primers / Q000235:genetics; D000070818:Discoidin Domain Receptors; D004392:Dwarfism / Q000000981:diagnostic imaging / Q000235:genetics*; D006801:Humans; D007150:Immunohistochemistry; D018613:Microscopy, Confocal; D008969:Molecular Sequence Data; D020125:Mutation, Missense / Q000235:genetics; D009896:Optic Atrophy / Q000473:pathology; D010009:Osteochondrodysplasias / Q000000981:diagnostic imaging / Q000235:genetics*; D010375:Pedigree; D010766:Phosphorylation; D021381:Protein Transport / Q000235:genetics; D011859:Radiography; D020794:Receptor Protein-Tyrosine Kinases / Q000235:genetics* / Q000378:metabolism*; D011975:Receptors, Mitogen / Q000235:genetics* / Q000378:metabolism*; D017422:Sequence Analysis, DNA; D017384:Sequence Deletion / Q000235:genetics; D035781:Siblings
chemical_list: D017931:DNA Primers; D011975:Receptors, Mitogen; D000070818:Discoidin Domain Receptors; D020794:Receptor Protein-Tyrosine Kinases
references.citation: Clin Dysmorphol. 1996 Jul;5(3):197-206
references.pmid: 8818447
references.citation: Physiol Genomics. 2009 Oct 7;39(2):120-9
references.pmid: 19671659
references.citation: Hum Mol Genet. 2009 Jan 15;18(2):267-77
references.pmid: 18945719
references.citation: Biochim Biophys Acta. 2013 Oct;1834(10):2187-94
references.pmid: 23128141
references.citation: Am J Hum Genet. 2009 Jan;84(1):80-4
references.pmid: 19110212
references.citation: EMBO Rep. 2001 May;2(5):446-52
references.pmid: 11375938
references.citation: Matrix Biol. 2006 Aug;25(6):355-64
references.pmid: 16806867
references.citation: J Biol Chem. 2002 Feb 1;277(5):3606-13
references.pmid: 11723120
references.citation: J Biol Chem. 2005 Jan 7;280(1):548-55
references.pmid: 15509586
references.citation: Am J Med Genet. 1993 Feb 1;45(3):320-6
references.pmid: 8434618
references.citation: Structure. 2009 Dec 9;17(12):1573-1581
references.pmid: 20004161
references.citation: Structure. 2012 Apr 4;20(4):688-97
references.pmid: 22483115
references.citation: Bone. 2014 Jan;58:33-41
references.pmid: 24120383
references.citation: Exp Eye Res. 2001 Jan;72(1):87-92
references.pmid: 11133186
references.citation: J Mol Biol. 2010 Jan 22;395(3):533-43
references.pmid: 19900459
references.citation: Clin Dysmorphol. 2009 Jan;18(1):31-35
references.pmid: 19050402
references.citation: Physiol Rev. 2012 Apr;92(2):537-76
references.pmid: 22535891
references.citation: J Biol Chem. 2003 May 9;278(19):16761-9
references.pmid: 12611880
references.citation: Hum Genet. 2007 Nov;122(3-4):389-95
references.pmid: 17665217
references.citation: J Bone Miner Res. 2011 Mar;26(3):604-17
references.pmid: 20734453
references.citation: Clin Dysmorphol. 2009 Jan;18(1):25-29
references.pmid: 19050401
references.citation: Nat Genet. 2011 Mar 06;43(4):306-8
references.pmid: 21378989
references.citation: Bioessays. 2003 Sep;25(9):868-77
references.pmid: 12938176
references.citation: Mol Reprod Dev. 2010 Jan;77(1):29-37
references.pmid: 19681157
references.citation: Biosci Biotechnol Biochem. 2013;77(9):1907-16
references.pmid: 24018687
references.citation: J Mol Biol. 2009 Jan 16;385(2):432-45
references.pmid: 19007791
references.citation: Cell. 2010 Jun 25;141(7):1117-34
references.pmid: 20602996
references.citation: Mol Cell. 1997 Dec;1(1):25-34
references.pmid: 9659900
references.citation: J Biol Chem. 2008 Mar 14;283(11):6861-8
references.pmid: 18201965
references.citation: J Bone Miner Res. 2010 Nov;25(11):2489-503
references.pmid: 20564243
references.citation: Hum Mol Genet. 2010 Jun 1;19(11):2239-50
references.pmid: 20223752
references.citation: Cancer Metastasis Rev. 2012 Jun;31(1-2):295-321
references.pmid: 22366781
references.citation: Mol Cell. 1997 Dec;1(1):13-23
references.pmid: 9659899
references.citation: Annu Rev Cell Dev Biol. 2011;27:265-90
references.pmid: 21568710
references.citation: FASEB J. 2001 May;15(7):1321-3
references.pmid: 11344127
references.citation: Med Hypotheses. 2009 Jan;72(1):55-7
references.pmid: 18851900
references.citation: Mol Endocrinol. 2008 Aug;22(8):1866-80
references.pmid: 18483174
references.citation: Traffic. 2000 Nov;1(11):836-51
references.pmid: 11208074
references.citation: Cell Signal. 2006 Aug;18(8):1108-16
references.pmid: 16626936
references.citation: Am J Med Genet. 1993 Feb 15;45(4):488-500
references.pmid: 8465857
references.citation: Dis Model Mech. 2013 Nov;6(6):1414-25
references.pmid: 24046357
references.citation: Int Rev Cell Mol Biol. 2014;310:39-87
references.pmid: 24725424
references.citation: Hum Mol Genet. 2005 Sep 1;14(17):2559-69
references.pmid: 16049033
references.citation: Nat Genet. 2004 Aug;36(8):801-8
references.pmid: 15284851
references.citation: J Biol Chem. 2006 Aug 11;281(32):22744-51
references.pmid: 16774916
references.citation: Traffic. 2002 Nov;3(11):781-90
references.pmid: 12383344