Localization of Fe(2+) at an RTGR sequence within a DNA duplex explains preferential cleavage by Fe(2+) and H2O2

Nicking of duplex DNA by the iron-mediated Fenton reaction occurs preferentially at a limited number of sequences. Of these, purine-T-G-purine (RTGR) is of particular interest because it is a required element in the upstream regulatory regions of many genes involved in iron and oxidative-stress responses. In order to study the basis of this preferential nicking, NMR studies were undertaken on the RTGR-containing duplex oligonucleotide, d(CGCGATATGACACTAG)/d(CTAGTGTCATATCGCG). One-dimensional and two-dimensional 1H NMR measurements show that Fe(2+) interacts preferentially and reversibly at the ATGA site within the duplex at a rate that is rapid relative to the chemical-shift timescale, while selective paramagnetic NMR line-broadening of the ATGA guanine H8 suggests that Fe(2+) interacts with the guanine N7 moiety. Localization at this site is supported by Fe(2+) titrations of a duplex containing a 7-deazaguanine substitution in place of the guanine in the ATGA sequence. The addition of a 100-fold excess of Mg(2+) over Fe(2+) does not affect the Fe(2+)-dependent broadening. When the ATGA site in the duplex is replaced by ATGT, an RTGR site (GTGA) is created on the opposite strand. Preferential iron localization then takes place at the 3' guanine in GTGA but no longer at the guanine in ATGT, consistent with the lack of preferential cleavage of ATGT sites relative to ATGA sites.     

Dehydroaltenusin, a mammalian DNA polymerase alpha inhibitor

Dehydroaltenusin was found to be an inhibitor of mammalian DNA polymerase alpha (pol alpha) in vitro. Surprisingly, among the polymerases and DNA metabolic enzymes tested, dehydroaltenusin inhibited only mammalian pol alpha. Dehydroaltenusin did not influence the activities of the other replicative DNA polymerases, such as delta and epsilon; it also showed no effect even on the pol alpha activity from another vertebrate (fish) or plant species. The inhibitory effect of dehydroaltenusin on mammalian pol alpha was dose-dependent, and 50% inhibition was observed at a concentration of 0.5 microm. Dehydroaltenusin-induced inhibition of mammalian pol alpha activity was competitive with the template-primer and non-competitive with the dNTP substrate. BIAcore analysis demonstrated that dehydroaltenusin bound to the core domain of the largest subunit, p180, of mouse pol alpha, which has catalytic activity, but did not bind to the smallest subunit or the DNA primase p46 of mouse pol alpha. These results suggest that the dehydroaltenusin molecule competes with the template-primer molecule on its binding site of the catalytic domain of mammalian pol alpha, binds to the site, and simultaneously disturbs dNTP substrate incorporation into the template-primer.     

A QUANTITATIVE STUDY OF TISSUE DYNAMICS DURING CLOSURE IN THE TRAPS OF VENUS'S FLYTRAP DIONAEA MUSCIPULA ELLIS

The mechanisms by which Venus's Flytrap (Dionaea muscipula Ellis) close are not clearly understood, and several conflicting models have been proposed. We have measured the dynamics of five trap tissues from three trap regions during full closure of young, fully developed, previously unclosed traps. Closure was divided into three distinct stages: 1) Capture–occurred immediately after stimulation of the trigger hairs and involved the rapid inward flexure of the trap margin and tynes. This motion interlocked the tynes, effectively capturing the prey. This was the only rapid movement of the trap; 2) Appression–completed by 30 min poststimulation, was characterized by contact of the margins; and 3) Sealing–completed by 1 hr poststimulation, was characterized by a sealed “digestive” sac formed around the potential prey, also by tight appression and recurved bending of the trap–margins. Major tissue dynamics that facilitated changes in trap morphology (hence, closure) occurred in different regions of the trap during different periods of time. The first regions where activity occurred were the A and C regions (Fig. 1), after approximately 15 min poststimulation; tissues in the C regions were most active followed by those in the B region of the trap (30 min to 1 hr poststimulation). Thus, shape changes during each stage of closure were the result of temporally separated changes in trap tissue volume. The complete sequence of events was elicited by a single 5–sec period of trigger hair stimulation. Our study showed that changes in the curvature of the trap during closure involved the expansion of opposing tissue groups (i.e., on opposite sides of trap medullary tissues). The pressure from contact of opposing trap lobes during the Appression stage may play an important role in regulating further trap closure and morphology.     

Inferences of genetic architecture of bill morphology in house sparrow using a high‐density SNP array point to a polygenic basis

Understanding the genetic architecture of quantitative traits can provide insights into the mechanisms driving phenotypic evolution. Bill morphology is an ecologically important and phenotypically variable trait, which is highly heritable and closely linked to individual fitness. Thus, bill morphology traits are suitable candidates for gene mapping analyses. Previous studies have revealed several genes that may influence bill morphology, but the similarity of gene and allele effects between species and populations is unknown. Here, we develop a custom 200K SNP array and use it to examine the genetic basis of bill morphology in 1857 house sparrow individuals from a large‐scale, island metapopulation off the coast of Northern Norway. We found high genomic heritabilities for bill depth and length, which were comparable with previous pedigree estimates. Candidate gene and genomewide association analyses yielded six significant loci, four of which have previously been associated with craniofacial development. Three of these loci are involved in bone morphogenic protein (BMP) signalling, suggesting a role for BMP genes in regulating bill morphology. However, these loci individually explain a small amount of variance. In combination with results from genome partitioning analyses, this indicates that bill morphology is a polygenic trait. Any studies of eco‐evolutionary processes in bill morphology are therefore dependent on methods that can accommodate polygenic inheritance of the phenotype and molecular‐scale evolution of genetic architecture.