Visualizing Rare Watson-Crick-Like Tautomeric and Anionic Mismatches in DNA and RNA

The central dogma of molecular biology relies on the correct Watson-Crick (WC) geometry of canonical deoxyribonucleic acid (DNA) dG•dC and dA•dT base pairs to replicate and transcribe genetic information with speed and an astonishing level of fidelity. In addition, the Watson-Crick geometry of canonical ribonucleic acid (RNA) rG•rC and rA•rU base pairs is highly conserved to ensure that proteins are translated with high fidelity. However, numerous other potential nucleobase tautomeric and ionic configurations are possible that can give rise to entirely new pairing modes between the nucleotide bases. Very early on, James Watson and Francis Crick recognized their importance and in 1953 postulated that if bases adopted one of their less energetically disfavored tautomeric forms (and later ionic forms) during replication it could lead to the formation of a mismatch with a Watson-Crick-like geometry and could give rise to “natural mutations.”

Since this time numerous studies have provided evidence in support of this hypothesis and have expanded upon it; computational studies have addressed the energetic feasibilities of different nucleobases’ tautomeric and ionic forms in siico; crystallographic studies have trapped different mismatches with WC-like geometries in polymerase or ribosome active sites. However, no direct evidence has been given for (i) the direct existence of these WC-like mismatches in canonical DNA duplex, RNA duplexes, or non-coding RNAs; (ii) which, if any, tautomeric or ionic form stabilizes the WC-like geometry. This thesis utilizes nuclear magnetic resonance (NMR) spectroscopy and rotating frame relaxation dispersion (R1ρ RD) in combination with density functional theory (DFT), biochemical assays, and targeted chemical perturbations to show that (i) dG•dT mismatches in DNA duplexes, as well as rG•rU mismatches RNA duplexes and non-coding RNAs, transiently adopt a WC-like geometry that is stabilized by (ii) an interconnected network of rapidly interconverting rare tautomers and anionic bases. These results support Watson and Crick’s tautomer hypothesis, but additionally support subsequent hypotheses invoking anionic mismatches and ultimately tie them together. This dissertation shows that a common mismatch can adopt a Watson-Crick-like geometry globally, in both DNA and RNA, and whose geometry is stabilized by a kinetically linked network of rare tautomeric and anionic bases. The studies herein also provide compelling evidence for their involvement in spontaneous replication and translation errors.

Serotonin transporter genotype modulates social reward and punishment in rhesus macaques

Background: Serotonin signaling influences social behavior in both human and nonhuman primates. In humans, variation upstream of the promoter region of the serotonin transporter gene (5-HTTLPR) has recently been shown to influence both behavioral measures of social anxiety and amygdala response to social threats. Here we show that length polymorphisms in 5-HTTLPR predict social reward and punishment in rhesus macaques, a species in which 5-HTTLPR variation is analogous to that of humans. Methodology/Principal Findings: In contrast to monkeys with two copies of the long allele (L/L), monkeys with one copy of the short allele of this gene (S/L) spent less time gazing at face than non-face images, less time looking in the eye region of faces, and had larger pupil diameters when gazing at photos of a high versus low status male macaques. Moreover, in a novel primed gambling task, presentation of photos of high status male macaques promoted risk-aversion in S/L monkeys but promoted risk-seeking in L/L monkeys. Finally, as measured by a "pay-per-view" task, S/L monkeys required juice payment to view photos of high status males, whereas L/L monkeys sacrificed fluid to see the same photos. Conclusions/Significance: These data indicate that genetic variation in serotonin function contributes to social reward and punishment in rhesus macaques, and thus shapes social behavior in humans and rhesus macaques alike. © 2009 Watson et al.

The neural basis of naming impairments in Alzheimer's disease revealed through positron emission tomography.

The naming impairments in Alzheimer's disease (AD) have been attributed to a variety of cognitive processing deficits, including impairments in semantic memory, visual perception, and lexical access. To further understand the underlying biological basis of the naming failures in AD, the present investigation examined the relationship of various classes of naming errors to regional brain measures of cerebral glucose metabolism as measured with 18 F-Fluoro-2-deoxyglucose (FDG) and positron emission tomography (PET). Errors committed on a visual naming test were categorized according to a cognitive processing schema and then examined in relationship to metabolism within specific brain regions. The results revealed an association of semantic errors with glucose metabolism in the frontal and temporal regions. Language access errors, such as circumlocutions, and word blocking nonresponses were associated with decreased metabolism in areas within the left hemisphere. Visuoperceptive errors were related to right inferior parietal metabolic function. The findings suggest that specific brain areas mediate the perceptual, semantic, and lexical processing demands of visual naming and that visual naming problems in dementia are related to dysfunction in specific neural circuits.

Spatial imagery preserves temporal order.

Line drawings were presented in either a spatial or a nonspatial format. Subjects recalled each of four sets of 24 items in serial order. Amount recalled in the correct serial order and sequencing errors were scored. In Experiment 1 items appeared either in consecutive locations of a matrix or in one central location. Subjects who saw the items in different locations made fewer sequencing errors than those who saw each item in a central location, but serial recall levels for these two conditions did not differ. When items appeared in nonconsecutive locations in Experiment 2, the advantage of the spatial presentation on sequencing errors disappeared. Experiment 3 included conditions in which both the consecutive and nonconsecutive spatial formats were paired with retrieval cues that either did or did not indicate the sequence of locations in which the items had appeared. Spatial imagery aided sequencing when, and only when, the order of locations in which the stimuli appeared could be reconstructed at retrieval.

New insights into Hoogsteen base pairs in DNA duplexes from a structure-based survey.

Hoogsteen (HG) base pairs (bps) provide an alternative pairing geometry to Watson-Crick (WC) bps and can play unique functional roles in duplex DNA. Here, we use structural features unique to HG bps (syn purine base, HG hydrogen bonds and constricted C1'-C1' distance across the bp) to search for HG bps in X-ray structures of DNA duplexes in the Protein Data Bank. The survey identifies 106 A•T and 34 G•C HG bps in DNA duplexes, many of which are undocumented in the literature. It also uncovers HG-like bps with syn purines lacking HG hydrogen bonds or constricted C1'-C1' distances that are analogous to conformations that have been proposed to populate the WC-to-HG transition pathway. The survey reveals HG preferences similar to those observed for transient HG bps in solution by nuclear magnetic resonance, including stronger preferences for A•T versus G•C bps, TA versus GG steps, and also suggests enrichment at terminal ends with a preference for 5'-purine. HG bps induce small local perturbations in neighboring bps and, surprisingly, a small but significant degree of DNA bending (∼14°) directed toward the major groove. The survey provides insights into the preferences and structural consequences of HG bps in duplex DNA.

Clinician judgment vs formal scales for predicting intracerebral hemorrhage outcomes.

OBJECTIVE: To compare the performance of formal prognostic instruments vs subjective clinical judgment with regards to predicting functional outcome in patients with spontaneous intracerebral hemorrhage (ICH). METHODS: This prospective observational study enrolled 121 ICH patients hospitalized at 5 US tertiary care centers. Within 24 hours of each patient's admission to the hospital, one physician and one nurse on each patient's clinical team were each asked to predict the patient's modified Rankin Scale (mRS) score at 3 months and to indicate whether he or she would recommend comfort measures. The admission ICH score and FUNC score, 2 prognostic scales selected for their common use in neurologic practice, were calculated for each patient. Spearman rank correlation coefficients (r) with respect to patients' actual 3-month mRS for the physician and nursing predictions were compared against the same correlation coefficients for the ICH score and FUNC score. RESULTS: The absolute value of the correlation coefficient for physician predictions with respect to actual outcome (0.75) was higher than that of either the ICH score (0.62, p = 0.057) or the FUNC score (0.56, p = 0.01). The nursing predictions of outcome (r = 0.72) also trended towards an accuracy advantage over the ICH score (p = 0.09) and FUNC score (p = 0.03). In an analysis that excluded patients for whom comfort care was recommended, the 65 available attending physician predictions retained greater accuracy (r = 0.73) than either the ICH score (r = 0.50, p = 0.02) or the FUNC score (r = 0.42, p = 0.004). CONCLUSIONS: Early subjective clinical judgment of physicians correlates more closely with 3-month outcome after ICH than prognostic scales.