Werner syndrome is a hereditary premature aging disorder characterized by genome instability. The product of the gene defective in WS, WRN, is a helicase/exonuclease that presumably functions in DNA metabolism. To understand the DNA structures WRN acts upon in vivo, we examined its substrate preferences for unwinding. WRN unwound a 3'-single-stranded (ss)DNA-tailed duplex substrate with streptavidin bound to the end of the 3'-ssDNA tail, suggesting that WRN does not require a free DNA end to unwind the duplex; however, WRN was completely blocked by streptavidin bound to the 3'-ssDNA tail 6 nucleotides upstream of the single-stranded/double-stranded DNA junction. WRN efficiently unwound the forked duplex with streptavidin bound just upstream of the junction, suggesting that WRN recognizes elements of the fork structure to initiate unwinding. WRN unwound two important intermediates of replication/repair, a 5'-ssDNA flap substrate and a synthetic replication fork. WRN was able to translocate on the lagging strand of the synthetic replication fork to unwind duplex ahead of the fork. For the 5'-flap structure, WRN specifically displaced the 5'-flap oligonucleotide, suggesting a role of WRN in Okazaki fragment processing. The ability of WRN to target DNA replication/repair intermediates may be relevant to its role in genome stability maintenance.

The brain-derived neurotrophic factor (BDNF) gene is critical for neuronal function and survival, and is likely to be important in psychiatric disorders. In this study, we used single-nucleotide polymorphism (SNP) discovery, functional analyses, and genetic association studies to better understand the potential role of BDNF sequence variation in behavior. Screening 480 unrelated individuals for SNPs and genotyping was performed in US Caucasian, American Indian, and African American populations. Lifetime DSM-III-R psychiatric diagnoses were assigned and the Tridimensional Personality Questionnaire (TPQ) was administered to measure anxious temperament (harm avoidance (HA)) and novelty seeking (NS). A novel SNP (À281 C4A) in promoter 1 was discovered that had decreased DNA binding in vitro and decreased basal reporter gene activity in transfected rat hippocampal neurons. The frequency of the À281 A allele was 0.03 in a Caucasian sample, but was virtually absent in other populations. Association analyses in a community-based sample showed that individuals with the À281 A allele (13 heterozygotes) had lower TPQ HA (F ¼ 4.8, po0.05). In contrast, the Met 66 allele was associated with increased HA (F ¼ 4.1, p ¼ 0.02) and was most abundant in individuals with both anxiety disorders and major depression (po0.05). Among the Val66Val homozygotes, individuals who were -281 CA heterozygotes had significantly lower HA than the -281 CC homozygotes (po0.01). Our results suggest that in this population, the low activity -281 A allele may be protective against anxiety and psychiatric morbidity, whereas Met 66 may be a risk allele.

Plains Indians have different smoking and drinking patterns and considerably less comorbidity between alcoholism and heavy smoking compared with the general U.S. population. Our COMT Val158Met results suggest that there may be both sex differences in the genetic origins of alcoholism and smoking in this population and overlap in genetic vulnerability to both addictions in women.