SNP Identification in α_(2A)-Adrenergic Receptor Gene in Chinese and the Effect on Gene Expression

Objective: To scan single nucleotide polymorphism ( SNP ) in Chinese alpha-2Aadrenergic receptor (α_(2A)-AR) gene and study the effects of the SNP on the gene expression.Methods: The complete sequence of α_(2A)-AR gene was analyzed with automated DNA sequencer to scanSNPs. Genomic DNA was extracted from whole blood and a 239 bp fragment containing the G/Cpolymorphism was amplified with PCR using a pair of. specific primers. PCR-RFLP was used to performthe genotyping of the SNP at the site-1 296 bp of the people in the North of China. Electrophoresismobility shift assay ( EMSA ) was used to study the binding of the 390 bp fragments (- 1 414-1 025bp) with G or C at the site-1 296 bp and nuclear extracts . Results: In our study, two SNPs werefound in α_(2A)-AR gene. Allele frequencies of the SNP at the site-1 296 bp were 0.61 and 0.39 forG and C , and the genotype frequencies were 0.34 , 0.54 and 0.13 for GG, GC and CC respectively fromthe people in the North of China. In the EMSA, a specific binding appeared in the complex ofnuclear extracts and DNA with C at-1 296 bp . Conclusion: Two SNPs exist in α_(2A)-AR gene from thepeople in the North of China , and DNA fragment with allele C of the SNP at the site-1 296 bp couldbind with a specific protein, which could influence the gene expression.

Genetic variations of beta 2-adrenergic receptor gene are associated with essential hypertension in Xinjiang Kazakans

Objective The aims of the present study were to investigate the associations of 46 A〉G, 79 C〉G, 491 C〉T and 659 C〉G genetic variants of the human beta 2-adrenergic receptor （β2-AR）, ADRB2, gene with essential hypertension （EH） in Xinjiang Kazakans population.Methods A gender-matched case-control （271 hypertensive cases and 267 normotensive controls） study was used to investigate the associations of the four variations in the coding region of ADRB2 with EH. The genotypes of the variants were identified by the polymerase chain reaction and restriction fragment length polymorphism （PCR-RFLP） methods. Results 46 A〉G, 79 C〉G and 659 C〉G polymorphisms were common in the Kazakan population, but 491 C〉T was a mutation （frequency ofT allele was only 0.003） and only found in EH group. The fxequency distributions of genotypes and alleles for 659 C〉G between the EH and control groups was significantly different （P〈0.05）, while those for 46 A〉G and 79 C〉G polymorphisms were not statistically different. Logistic regression analysis suggested that the G allele of 659 C〉G polymorphism was a risk factor for hypertension （minor allele vs common homo; odds ratio, 13.240, 95% CI, 4.052-43.274; P〈0.05）. Covariance analysis showed that systolic and diastolic blood pressure levels in GG＋CG group of 659 C〉G were significantly higher than those in the CC group, but no significant difference of blood pressure were found between common homo and minor allele for 46 A〉G and 79C〉G polymorphisms. Haplotype analysis showed that two hyplotypes, HI： 46A-79C-491C-523C（48%）and H5：46A-79C-491C-659G, were associated with EH.Conelusion ADRB2 genetic variants may play independent roles in the molecular genetic mechanism of EH in Xinjiang Kazakans population （d Geriatr Cardio12010; 7：52-57）.

Lack of association between ADRA2B-4825 gene insertion/deletion poly- morphism and migraine in Chinese Han population

Objective The present study aimed to estimate the association between susceptibility to migraine and the 12nucleotide insertion/deletion （indel） polymorphism in promoter region ofα 2B -adrenergic receptor gene （ADRA2B）.Methods A case-control study was carried out in Chinese Han population,including 368 cases of migraine and 517 controls.Genomic DNA was extracted from blood samples,and DNA fragments containing the site of polymorphism were amplified by PCR.Data were adjusted for sex,age,migraine history and family history,and analyzed using a logistic regression model.Results There was no association between indel polymorphism and migraine,at either the allele or the genotype level.Conclusion These findings do not support a functional significance of ADRA2B indel polymorphism at position-4825 relative to the start codon in the far upstream region of the promoter in the present migraine subjects.

A novel β_(2)-AR agonist,Higenamine,induces β-arrestin-biased signaling

The biased ligands in G protein-coupled receptors(GPCRs)have opened new avenues for developing safer and more effective drugs.However,the identification of such biased ligands as drug candidates is highly desirable.Here,we report that Higenamine,a compound isolated from a Chinese herb,functions as a novel β-arrestin-biased ligand of the β_(2)-adrenergic receptor(β_(2)-AR).The radioligand binding assays demonstrated that Higenamine was the ligand of β_(2)-AR.Higenamine induced phosphorylation of extracellular signal-regulated kinase 1/2(ERK1/2),which can be blocked by propranolol,an inhibitor of β_(2)-AR.The Gi protein inhibitor,pertussis toxin,had no effect on the phosphorylation of ERK1/2 induced by Higenamine.Furthermore,Higenamine induced ERK1/2 phosphorylation through transactivation of Epithelial growth factor receptor(EGFR).We also found that Higenamine-induced-ERK1/2 phosphorylation is dependent on β-arrestin1/2,and HG inhibits Doxorubicin-induced cardiomyocyte apoptosis.Our results identify Higenamine as a novel biased ligand via the β-arrestin-dependent pathway.These findings give us a better understanding of Higenamine’s potential role in designing diagnostic and therapeutic strategies.