To identify potentially functional sequence variants in the 5 region of NTRK2, 3.4 kb of the gene was analyzed in 190 unrelated individuals. The genomic region screened included five known exons in the 5 untranslated region (5-UTR) and flanking sequence, comprising 3.1 kb of genomic DNA relative to the translation start site. The resequencing sample was composed of 40 African Americans, 40 Finnish Caucasians, and 110 U.S. Caucasians. The sample was also clinically diverse. The screening strategy used high performance liquid chromatography (dHPLC). PCR samples with differential dHPLC elution profiles were selected for direct sequence analysis.? For association studies, a total of 43 loci from NTRK2 were selected for genotyping. The markers covered the entire NTRK2 region with a mean distance of 8796 kb ( 5992 kb) between markers. Genotyping was performed by using 5- nuclease assays (TaqMan, Applied Biosystems, Inc.; ABI). Error rates were determined from genotypes of from 10% of the samples, taken at random. The sample Finnish Caucasian population was composed of 516 psychiatrically interviewed subjects who included 229 subjects with alcohol dependence (AD) and 287 healthy controls. ? Resequencing of the 5 region of NTRK2 uncovered two novel sequence variants (designated LNG5 and LNG7). The two previously unknown SNPs were located within intronic sequences. Five of the seven variants were previously known and were located in either the 5 flanking region or 5-UTR. These other five SNPs were reported in the National Center for Biotechnology Information (NCBI) Single Nucleotide Polymorphism Database (dbSNP) or the Celera Discovery Systems databases. Celera database and NCBI entries are available through the NCBI and the HapMap Project. Preliminary allele frequencies for each SNP were determined from the number of heterozygotes detected by denaturing high-performance liquid chromatography in the screening panel. Variants LNG5 and LNG7 each were detected at an estimated allele frequency of 0.03. This was a preliminary allele frequency based on the entire screening panel, which was composed of individuals from different ethnic populations. Because SNPs LNG5 and LNG7 were previously unknown, we incorporated them into the marker list. ? Genotypes were obtained from a total of 43 SNP markers spanning 363 kb of the NTRK2 gene and were tested for meeting Hardy-Weinberg equilibrium (HWE) expectations in the case and control groups, respectively. All SNPs fit HWE in each group (P > 0.05) with the exception of SNP 19 (P = 0.042) and SNP 32, (P = 0.045), which showed marginally significant departures from HWE. These two markers were excluded from subsequent analyses.? The remaining SNPs were analyzed for allelic and genotypic association. For two markers, strong significant allelic association with AD was observed (P = 0.0041 for SNP5 and P = 0.0019 for SNP30). SNP 4 and SNP 30 showed genotypic association with AD after Bonferroni correction (P = 0.01). Of note was the fact that the three significant SNPs (4, 5, and 30) were represented more frequently in the AD group than in the controls, suggesting that these SNPs were associated with AD in the Finnish population.? We then determined pairwise D-prime values across the 43 markers in the Finnish control population. Overall linkage disequilibrium (LD) was strong across the 353 kb region. Five LD blocks were observed: block 1, located at the 5 end of NTRK2 and exons 1 through 15 which encode the extracellular and transmembrane domains) covering a physical region of at least 119 kb, accounting for nearly 34% of NTRK2. Within block 1, more than 85% of SNP pairs showed complete LD (D>0.99). ? In LD block1, eighteen loci generated five haplotypes with frequencies greater than 0.03 and represented 91% of chromosomes in the control group and 86% in the AD group. The global haplotype comparison in block 1 showed a marginally significant difference (global P = 0.057). For a single haplotype comparison between control and AD groups, haplotype 1A (which embraced the major allele of SNP4 and SNP5) was represented more frequently in the unaffected control group (0.604) than in the AD group (0.527) (Chi square = 5.898, P = 0.013). The P- value remained marginally significant after correction for multiple tests (P = 0.065). No other individual haplotypes from the other four blocks were significantly associated with AD.? This was the first report of NTRK2 association with AD. The data suggest that variation at the NTRK2 gene may be protective in AD. Although our initial resequencing effort was confined to the 5 flanking region, it uncovered two previously unknown SNPs. One of these sequence variants (SNP2) is located between two alternative 5 UTR exons while the other (SNP4) is located within intron 5, which is 3 of the first coding exon. Their functional impact on NTRK2 regulation is not known.? In a second study, we determined that a marker in the GRIK4 gene, which encodes the KA1, kainic acid type ionotropic glutmate receptor, met prior P-value thresholds for treatment response to the SSRI citalopram. The study population comprised adults with recurrent major depression who participated in the Sequenced Treatment Alternatives to Relieve Depression (STAR*D) trial (www.star-d.org). DNA was collected from 1953 participants from this study who received the selective serotonin reuptake inhibitor (SSRI) citalopram in primary and psychiatric care settings followed by regular assessment of outcome and side effects. Homozygous carriers of response-associated markers from both GRIK4 and a previously associated HTR2A marker were 23% less likely to experience non-response to treatment as patients carrying none of these marker alleles. The findings suggest a role for the glutamatergic neurotransmitter system in modulating response to the SSRI citalopram
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