Characterization of the mechanical unfolding of RNA pseudoknots

The pseudoknot is an important RNA structural element that provides an excellent model system for studying the contributions of tertiary interactions to RNA stability and to folding kinetics. RNA pseudoknots are also of interest because of their key role in the control of ribosomal frameshifting by viral RNAs. Their mechanical properties are directly relevant to their unfolding by ribosomes during translation. We have used optical tweezers to study the kinetics and thermodynamics of mechanical unfolding and refolding of single RNA molecules. Here we describe the unfolding of the frameshifting pseudoknot from infectious bronchitis virus (IBV), three constituent hairpins, and three mutants of the IBV pseudoknot. All four pseudoknots cause −1 programmed ribosomal frameshifting. We have measured the free energies and rates of mechanical unfolding and refolding of the four frameshifting pseudoknots. Our results show that the IBV pseudoknot requires a higher force than its corresponding hairpins to unfold. Furthermore, its rate of unfolding changes little with increasing force, in contrast with the rate of hairpin unfolding. The presence of Mg²⁺ significantly increases the kinetic barriers to unfolding the IBV pseudoknot, but has only a minor effect on the hairpin unfolding. The greater mechanical stability of pseudoknots compared to hairpins, and their kinetic insensitivity to force supports the hypothesis that −1 frameshifting depends on the difficulty of unfolding the mRNA.     

Laurus novocanariensis essential oil: Seasonal variation and valorization

We investigated the qualitative and quantitative seasonal variation of the leaf and fruit oils of the Macaronesian endemism Laurus novocanariensis and their plant defensive potential. The monoterpene fraction dominated the leaf (74%) and berry essential oils (73–44%, ripe–unripe). The insect antifeedant effects of these oils were species- and season-dependent against the aphids (Myzus persicae and Rhopalosiphum padi). Overall, the biological effects of these oils correlated with the oxygenated terpene fraction. Among the pure components tested, β-caryophyllene and its oxide were strong antifeedants to Leptinotarsa decemlineata and Spodoptera littoralis. The aphids responded to β-ocimene, β-pinene, 1,8-cineole, linalool (antifeedants) and linalool oxide (attractive to M. persicae). The antifungal effects of the leaf oils on Fusarium spp. were season-dependent. β-Caryophyllene oxide proved to be a strong antifungal. L. novocanariensis oils inhibited Lactuca sativa germination and radicle elongation, the leaves being more effective. Linalool also inhibited seed germination.     

Polynucleotide circular dichroism calculations: use of an all-order classical coupled oscillator polarizability theory.

An all-order classical coupled oscillator theory in which monomer band shapes are explicity taken into account is applied to the calculations of polynucleotide circular dichroism. Calculations are shown for ApA and oligoadenylic acid of varying chain lengths, in both RNA and B-DNA geometry, and the advantages of this theory are assessed. By introducing an effective dielectric constant, good agreement with measured spectra is obtained. Variations in monomer parameters are tested in an attempt to eliminate remaining discrepancies between calculated and measured spectra.     

Hepatocellular carcinoma in two captive golden‐headed lion tamarins (Leontopithecus chrysomelas)

Two cases of hepatocellular carcinoma (HCC) in 2 female captive golden‐headed lion tamarins (Leontopithecus chrysomelas) are described. HCC was diagnosed in both, with pulmonary metastasis in one of them. Neoplastic cells were positive for hepatocyte‐specific antigen (HSA) by immunohistochemistry, confirming the diagnosis.     

Utilization of C20-polyunsaturated fatty acids by a yeast fatty acid desaturase mutant

A study was made of the utilization of C₂₀-polyunsaturated fatty acids by the $\text{S. cerevisiae}$ fatty acid desaturase mutant $\text{olel-1}$, Arachidonic acid, 8,11,14-eicosatrienoic acid, and 5,8,11,14,17-eicosapentaenoic acid were about equally effective in supporting growth with lactate as the carbon source. The relative proportion of these fatty acids in total cell fatty acids was $\text{ca}$. 50%. 5,8,11-eicosatrienoic acid synthesized from oleate was less effective. Very little growth occurred with 11,14,17-eicosatrienoic acid or with 11,14-eicosadienoic acid. These results indicate the usefulness of the yeast mutant as a eucaryotic model for study of membrane systems enriched in specific C₂₀-polyunsaturated fatty acids.     

Force unfolding kinetics of RNA using optical tweezers. I. Effects of experimental variables on measured results

Experimental variables of optical tweezers instrumentation that affect RNA folding/unfolding kinetics were investigated. A model RNA hairpin, P5ab, was attached to two micron-sized beads through hybrid RNA/DNA handles; one bead was trapped by dual-beam lasers and the other was held by a micropipette. Several experimental variables were changed while measuring the unfolding/refolding kinetics, including handle lengths, trap stiffness, and modes of force applied to the molecule. In constant-force mode where the tension applied to the RNA was maintained through feedback control, the measured rate coefficients varied within 40% when the handle lengths were changed by 10-fold (1.1-10.2 Kbp); they increased by two- to threefold when the trap stiffness was lowered to one-third (from 0.1 to 0.035 pN/nm). In the passive mode, without feedback control and where the force applied to the RNA varied in response to the end-to-end distance change of the tether, the RNA hopped between a high-force folded-state and a low-force unfolded-state. In this mode, the rates increased up to twofold with longer handles or softer traps. Overall, the measured rates remained with the same order-of-magnitude over the wide range of conditions studied. In the companion article on pages 3010-3021, we analyze how the measured kinetics parameters differ from the intrinsic molecular rates of the RNA, and thus how to obtain the molecular rates.     

Effects of hummingbird morphology on specialization in pollination networks vary with resource availability

Specialization of species in interaction networks influences network stability and ecosystem functioning. Spatial and temporal variation in resource availability may provide insight into how ecological factors, such as resource abundance, and evolutionary factors, such as phylogenetically conserved morphological traits, influence specialization within mutualistic networks. We used independent measures of hummingbird abundance and resources (nectar), information on hummingbird traits and plant–hummingbird interactions to examine how resource availability and species' morphology influence the specialization of hummingbirds in three habitat types (forest, shrubs, cattle ranch) sampled over 10 sessions across two years in the southern Andes of Ecuador. Specialization of hummingbird species in the networks was measured by three indices: d' (related to niche partitioning), generality (related to niche width) and PSI (related to pollination services). Specialization indices d', generality and PSI of hummingbird species were influenced by resource availability. All indices indicated that specialization of hummingbirds increased when the availability of resources decreased. Variation in d' was also explained by an interaction between resource availability and bill length; hummingbirds with a long bill switched from being more specialized than other species when resource availability was low to being similarly specialized when availability was high. Overall, our results highlight the importance of ecological and evolutionary factors determining the specialization of species in interaction networks. We demonstrate in particular that ecological gradients in resource availability cause substantial changes in consumers' foraging behavior contingent on their morphology. Changes in pollinator specialization along resource gradients can have impacts on ecosystem functions, such as pollination by animals.