Molecular scanning of the beta-3-adrenergic receptor gene in Pima Indians and Caucasians.
BACKGROUND
The beta-3-adrenergic receptor (beta3AR) stimulates lipolysis and thermogenesis in adipocytes. The Trp64Arg beta3AR variant is associated in some, but not all, studies with an earlier onset of Type 2 diabetes mellitus and features of the insulin resistance syndrome. Functional studies as to the role of the Trp64Arg variant have been inconclusive. Earlier studies screened the beta3AR gene in only ten obese, diabetic Pima Indians. Potentially another yet to be identified polymorphism in the beta3AR gene in linkage disequilibrium with the Trp64Arg polymorphism could explain the findings in the association and functional studies.


METHODS
We scanned the beta3AR gene in 20 diabetic Pima subjects and 20 Caucasian subjects using single stranded conformational polymorphism (SSCP) analysis. Variants were sequenced using dideoxy sequence analysis and further characterized using allele specific oligonucleotide hybridization (ASO) and RNA template specific-polymerase chain reaction (RS-PCR) assays.


RESULTS
We found a guanine to thymidine substitution in the first intron, 14 bases from the splice donor site in both groups. In virtually all subjects, only two haplotypes were detected, Trp64/g1856 and Arg64/t1856, indicating that the g1856t polymorphism is in linkage disequilibrium with the Trp64Arg polymorphism. The g1856t substitution introduces a new consensus splice donor site which, if used, would encode a truncated protein. RNA levels of the two beta3AR alleles were approximately equal in omental adipose tissue of heterozygotes. No aberrantly spliced beta3AR mRNA was detected, indicating that the new consensus splice donor site is not used in vivo.


CONCLUSIONS
The g1856t polymorphism is in linkage disequilibrium with the Trp64Arg variant, but does not appear to have a functional role.
Silver
K
K
Division of Endocrinology and Metabolism, Johns Hopkins University School of Medicine, Baltimore, MD 21201, USA. ksilver@umppa1.ab.umd.edu
Walston
J
J
Yang
Y
Y
Pratley
R
R
Ravussin
E
E
Raben
N
N
Shuldiner
A R
AR
D019943:Amino Acid Substitution / Q000235:genetics; D001120:Arginine / Q000235:genetics / Q000378:metabolism; D003924:Diabetes Mellitus, Type 2 / Q000235:genetics*; D005838:Genotype; D006801:Humans; D007198:Indians, North American / Q000235:genetics*; D009154:Mutation; D012150:Polymorphism, Restriction Fragment Length; D018807:Polymorphism, Single-Stranded Conformational; D012326:RNA Splicing; D012333:RNA, Messenger / Q000032:analysis / Q000235:genetics; D011943:Receptors, Adrenergic, beta / Q000235:genetics*; D022702:Receptors, Adrenergic, beta-3; D020133:Reverse Transcriptase Polymerase Chain Reaction; D017421:Sequence Analysis; D014364:Tryptophan / Q000235:genetics / Q000378:metabolism; D044465:White People / Q000235:genetics*
D012333:RNA, Messenger; D011943:Receptors, Adrenergic, beta; D022702:Receptors, Adrenergic, beta-3; D014364:Tryptophan; D001120:Arginine