title: Analysis of a rare functional truncating mutation rs61757459 in vaspin (SERPINA12) on circulating vaspin levels.
abstract: A recent genome-wide association study suggests that genetic variation within the vaspin gene might contribute to the variability in circulating serum visceral adipose tissue-derived serine protease inhibitor (vaspin) concentrations. Here, we analyzed the functional consequences of the rare variant rs61757459 predicting a premature stop codon and its impact on circulating serum vaspin concentrations. In order to identify genetic variation, we sequenced the vaspin gene in 48 nonrelated Caucasian subjects. Rs61757459 was subsequently genotyped in three metabolically well-characterized German cohorts (N = 4,019). We addressed the impact of rs61757459 on the crystal structure of vaspin and investigated its effects on vaspin expression in vivo as well as in vitro using various cell lines (Escherichia coli, HEK293). Along with previously reported common genetic variants, sequencing of vaspin revealed a rare variant (rs61757459; minor allele frequency: 1 %) which predicts a premature stop codon p.R211X. Heterozygous carriers of this mutation had lower circulating vaspin levels when compared with noncarriers. In silico structure analysis of the truncated vaspin, which was estimated to be 24.5 kDa, suggested misfolding and potential instability due to the absence of core structural domains. Indeed, the truncated protein was detected after recombinant expression in E. coli and in lysate, but not in supernatant of HEK293 cells. We conclude that rs61757459 is a functional mutation that results in a truncated protein whose instability likely results in reduced serum vaspin levels.
  
  
  CONCLUSIONS
  A rare variant (rs61757459) in vaspin coding for the stop codon p.R211X is related to lower circulating vaspin concentrations. Structure analysis suggests misfolding and instability due to the absence of core structural domains. The truncated protein is detectable after recombinant expression in E. coli and in lysate, but not in supernatant of HEK293 cells.
authors.affiliation: Department of Medicine, University of Leipzig, Leipzig, Germany.
authors.forename: Jana
authors.initials: J
authors.lastname: Breitfeld
authors.forename: John T
authors.initials: JT
authors.lastname: Heiker
authors.forename: Yvonne
authors.initials: Y
authors.lastname: Böttcher
authors.forename: Dorit
authors.initials: D
authors.lastname: Schleinitz
authors.forename: Anke
authors.initials: A
authors.lastname: Tönjes
authors.forename: Kerstin
authors.initials: K
authors.lastname: Weidle
authors.forename: Kerstin
authors.initials: K
authors.lastname: Krause
authors.forename: E Bartholomeus
authors.initials: EB
authors.lastname: Kuettner
authors.forename: Markus
authors.initials: M
authors.lastname: Scholz
authors.forename: Wieland
authors.initials: W
authors.lastname: Kiess
authors.forename: Norbert
authors.initials: N
authors.lastname: Sträter
authors.forename: Annette G
authors.initials: AG
authors.lastname: Beck-Sickinger
authors.forename: Michael
authors.initials: M
authors.lastname: Stumvoll
authors.forename: Antje
authors.initials: A
authors.lastname: Körner
authors.forename: Matthias
authors.initials: M
authors.lastname: Blüher
authors.forename: Peter
authors.initials: P
authors.lastname: Kovacs
mesh_terms: D000368:Aged; D018388:Codon, Terminator; D015331:Cohort Studies; D004926:Escherichia coli / Q000235:genetics; D005260:Female; D015870:Gene Expression; D057809:HEK293 Cells; D006801:Humans; D008297:Male; D008875:Middle Aged; D008958:Models, Molecular; D009154:Mutation; D010802:Phylogeny; D017510:Protein Folding; D055550:Protein Stability; D011994:Recombinant Proteins / Q000032:analysis / Q000235:genetics; D015843:Serpins / Q000032:analysis / Q000097:blood* / Q000235:genetics*
chemical_list: D018388:Codon, Terminator; D011994:Recombinant Proteins; C507339:SERPINA12 protein, human; D015843:Serpins