Several transporters belonging to the ABCA subfamily of ABC (ATP-binding cassette) proteins are involved in lipid trafficking. Human ABCA5 and its rat orthologue, rAbca5, represent recently identified subfamily members whose substrate spectrum remains to be defined. The elucidation of (sub)cellular rAbca5 distribution would be expected to provide a basis for optimization of functional analyses. In the present study, we applied in situ hybridization to examine rAbca5 mRNA distribution within sections of rat testis, a tissue expressing high levels of rAbca5 mRNA. We found rAbca5 mRNA to be predominantly expressed in interstitial Leydig cells, which are major sites of testosterone synthesis. To investigate rAbca5 subcellular localization, we constructed expression vectors yielding rAbca5 fused either to EGFP (enhanced green fluorescent protein) or to a peptide bearing the viral V5 epitope. During rAbca5 cDNA cloning, we discovered a splice variant sequence (rAbca5 V20+16), predicted to give rise to a truncated, half-size transporter, which was highly homologous with a human splice variant described by us previously. Quantitative RT (reverse transcription)-PCR demonstrated that the rAbca5 splice variant was expressed in numerous tissues (including testis, brain and lungs), its cDNA amounting to 2.6-11.2% of total rAbca5 cDNA. Transfection of individual rAbca5-EGFP, rAbca5 splice variant-EGFP or transporter-V5 expression plasmids along with organelle marker plasmids into HEK-293 cells (human embryonic kidney 293 cells) revealed that both rAbca5 and splice variant fusion proteins co-localized with marker protein for the Golgi apparatus. Expression of rAbca5 mRNA in Leydig cells, intracellular localization of rAbca5-EGFP/rAbca5-V5 and involvement of rAbca5-related proteins in lipid transport suggest that rAbca5 may participate in intracellular sterol/steroid trafficking.
The -100_-102delAAG 3 bp deletion increases the HTR3B promoter activity in vitro. The consequences of this for the structure and the function of the resulting 5-HT3 receptors remain to be elucidated.
Glucocorticoids seem to mediate the effect of stimulant drugs such as nicotine. Several studies have pointed to an association between the BclI polymorphism in the glucocorticoid receptor gene and increased glucocorticoid effects. We analysed the association of smoking behaviour and the BclI polymorphism using a case-control design within the framework of a larger pharmacogenetic study. A total of 327 Caucasian patients with asthma or chronic obstructive pulmonary disease from 39 German general practices gave informed consent to take part in the study. They filled in questionnaires concerning their smoking behaviour and were genotyped for the BclI polymorphism. The genotype frequencies for non-smokers (n = 251; CC, 0.42; CG, 0.46; GG, 0.12) as well as for smokers (n = 76; CC, 0.29; CG, 0.55; GG, 0.16) were consistent with the Hardy-Weinberg equilibrium. The proportion of smokers was significantly lower among carriers of the CC-genotype (22/127 = 17%) compared with carriers of the G-allele (54/200 = 27%; chi2 = 4.08; P = 0.04). Within the group of smokers, the proportion of heavy smokers (> 19 cigarettes/day; median) was reduced in C-homozygous patients when compared with carriers of the G-allele (7/22 = 32% versus 31/54 = 57%; chi2 = 4.09; P = 0.04). Stepwise logistic regression analysis also pointed to an association between the CC-genotype and a reduced probability of being a smoker (odds ratio = 0.55; 95% confidence interval = 0.30-1.00; P = 0.05) controlling for other predictors. In summary, this study provides evidence that the BclI polymorphism might play a role in the maintenance and severity of nicotine dependence.
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