Journals
2009 EN
Diane Martin · Omid Toub · Angela Chiang
+4 more
Terpenoid volatiles are important information molecules that enable pollinators to locate flowers and may protect reproductive tissues against pathogens or herbivores. Inflorescences of grapevine (Vitis vinifera L.) are composed of tiny green flowers that produce an abundance of sesquiterpenoid volatiles. We demonstrate that male flower parts of grapevines are responsible for sesquiterpenoid floral scent formation. We describe temporal and spatial patterns of biosynthesis and release of floral volatiles throughout the blooming of V. vinifera L. cv. Cabernet Sauvignon. The biosynthesis of sesquiterpene volatiles, which are emitted with a light-dependent diurnal pattern early in the morning at prebloom and bloom, is localized to anthers and, more specifically, within the developing pollen grains. Valencene synthase (VvValCS) enzyme activity, which produces the major sesquiterpene volatiles of grapevine flowers, is present in anthers. VvValCS transcripts are most abundant in flowers at prebloom stages. Western blot analysis identified VvValCS protein in anthers, and in situ immunolabeling located VvValCS protein in pollen grains during bloom. Histochemical staining, as well as immunolabeling analysis by fluorescent microscopy and transmission electron microscopy, indicated that VvValCS localizes close to lipid bodies within the maturing microspore.
National Academy of Sciences
Journals
2009 EN
Barry R. O’Keefe · Fakhrieh Vojdani · Viviana Buffa
+11 more
To prevent sexually transmitted HIV, the most desirable active ingredients of microbicides are antiretrovirals (ARVs) that directly target viral entry and avert infection at mucosal surfaces. However, most promising ARV entry inhibitors are biologicals, which are costly to manufacture and deliver to resource-poor areas where effective microbicides are urgently needed. Here, we report a manufacturing breakthrough for griffithsin (GRFT), one of the most potent HIV entry inhibitors. This red algal protein was produced in multigram quantities after extraction from Nicotiana benthamiana plants transduced with a tobacco mosaic virus vector expressing GRFT. Plant-produced GRFT (GRFT-P) was shown as active against HIV at picomolar concentrations, directly virucidal via binding to HIV envelope glycoproteins, and capable of blocking cell-to-cell HIV transmission. GRFT-P has broad-spectrum activity against HIV clades A, B, and C, with utility as a microbicide component for HIV prevention in established epidemics in sub-Saharan Africa, South Asia, China, and the industrialized West. Cognizant of the imperative that microbicides not induce epithelial damage or inflammatory responses, we also show that GRFT-P is nonirritating and noninflammatory in human cervical explants and in vivo in the rabbit vaginal irritation model. Moreover, GRFT-P is potently active in preventing infection of cervical explants by HIV-1 and has no mitogenic activity on cultured human lymphocytes.
National Academy of Sciences
Journals
2009 EN
Morgan W. Tingley · William B. Monahan · Steven R. Beissinger
+1 more
In the face of environmental change, species can evolve new physiological tolerances to cope with altered climatic conditions or move spatially to maintain existing physiological associations with particular climates that define each species' climatic niche. When environmental change occurs over short temporal and large spatial scales, vagile species are expected to move geographically by tracking their climatic niches through time. Here, we test for evidence of niche tracking in bird species of the Sierra Nevada mountains of California, focusing on 53 species resurveyed nearly a century apart at 82 sites on four elevational transects. Changes in climate and bird distributions resulted in focal species shifting their average climatological range over time. By comparing the directions of these shifts relative to the centroids of species' range-wide climatic niches, we found that 48 species (90.6%) tracked their climatic niche. Analysis of niche sensitivity on an independent set of occurrence data significantly predicted the temperature and precipitation gradients tracked by species. Furthermore, in 50 species (94.3%), site-specific occupancy models showed that the position of each site relative to the climatic niche centroid explained colonization and extinction probabilities better than a null model with constant probabilities. Combined, our results indicate that the factors limiting a bird species' range in the Sierra Nevada in the early 20th century also tended to drive changes in distribution over time, suggesting that climatic models derived from niche theory might be used successfully to forecast where and how to conserve species in the face of climate change.
National Academy of Sciences
Journals
2009 EN
Steven M. Abel · YingLung Steve Tse · Hans C. Andersenb
The one-dimensional, single-occupancy lattice gas exhibits highly cooperative particle motion and provides an interesting challenge for theoretical methods designed to describe caging in liquids. We employ this model in an effort to gain insight into caging phenomena in more realistic models of liquids, using a diagrammatic kinetic theory of density fluctuations to develop a series of approximations to the kinetic equations for the van Hove self-correlation function. The approximations are formulated in terms of the irreducible memory function, and we assess their efficacy by comparing their solutions with computer simulation results and the well-known subdiffusive behavior of a tagged particle at long times. The first approximation, a mode coupling theory, factorizes the 4-point propagators that contribute to the irreducible memory function into products of independent single-particle propagators. This approximation fails to capture the subdiffusive behavior of a tagged particle at long times. Analysis of the mode coupling approximation in terms of the diagrammatic kinetic theory leads to the development of two additional approximations that can be viewed as diagrammatic extensions or modifications of mode coupling theory. The first, denoted MC1, captures the long-time subdiffusive behavior of a tagged particle. The second, denoted MC2, captures the subdiffusive behavior of a tagged particle and also yields the correct amplitude of its mean square displacement at long times. Numerical and asymptotic solutions of the approximate kinetic equations share many qualitative and quantitative features with simulation results at all timescales.
National Academy of Sciences
Journals
2009 EN
P.V. Ganesan · Steven G. Boxer
Freestanding phospholipid bilayers have been assembled spanning shallow, micrometer-sized wells etched into a Si wafer substrate so that the bilayers are near (within hundreds of nanometers) but not in contact with the wafer surface. The proximity of the bilayers to the highly reflective Si/SiO(2) interface allows them to be probed by using fluorescence-interference techniques. These interferometry measurements show that the bilayers are curved and that the curvature can be varied by changes in osmotic pressure. Furthermore, the ionophore gramicidin can be incorporated into the bilayers, making them selectively permeable to monovalent cations. This freestanding architecture may overcome surface-interaction problems that occur when cell membrane proteins are introduced into solid supported bilayers, while also allowing for high-precision measurements of changes in fluorophore position by interferometry.
National Academy of Sciences
Journals
2009 EN
Jonathan F. Arambula · Sreenivasa Rao Ramisetty · Anne M. Baranger
+1 more
This work describes the rational design, synthesis, and study of a ligand that selectively complexes CUG repeats in RNA (and CTG repeats in DNA) with high nanomolar affinity. This sequence is considered a causative agent of myotonic dystrophy type 1 (DM1) because of its ability to sequester muscleblind-like (MBNL) proteins. Ligand 1 was synthesized in two steps from commercially available compounds, and its binding to CTG and CUG repeats in oligonucleotides studied. Isothermal titration calorimetry studies of 1 with various sequences showed a preference toward the T-T mismatch (K(d) of 390 +/- 80 nM) with a 13-, 169-, and 85-fold reduction in affinity toward single C-C, A-A, and G-G mismatches, respectively. Binding and Job analysis of 1 to multiple CTG step sequences revealed high affinity binding to every other T-T mismatch with negative cooperativity for proximal T-T mismatches. The affinity of 1 for a (CUG)(4) step provided a K(d) of 430 nM with a binding stoichiometry of 1:1. The preference for the U-U in RNA was maintained with a 6-, >143-, and >143-fold reduction in affinity toward single C-C, A-A, and G-G mismatches, respectively. Ligand 1 destabilized the complexes formed between MBNL1N and (CUG)(4) and (CUG)(12) with IC(50) values of 52 +/- 20 microM and 46 +/- 7 microM, respectively, and K(i) values of 6 +/- 2 microM and 7 +/- 1 microM, respectively. These values were only minimally altered by the addition of competitor tRNA. Ligand 1 does not destabilize the unrelated RNA-protein complexes the U1A-SL2 RNA complex and the Sex lethal-tra RNA complex. Thus, ligand 1 selectively destabilizes the MBNL1N-poly(CUG) complex.
National Academy of Sciences
Journals
2009 EN
Christopher Thaler · Srinagesh V. Koushik · Henry L. Puhl
+2 more
At its fundamental level, human memory is thought to occur at individual synaptic contact sites and manifest as persistent changes in synaptic efficacy. In digital electronics, the fundamental structure for implementing memory is the flip-flop switch, a circuit that can be triggered to flip between two stable states. Recently, crystals of Ca(2+)/calmodulin-dependent protein kinase IIalpha (CaMKIIalpha) catalytic domains, the enzymatic portion of a dodecameric holoenzyme involved in memory, were found to form dimers [Rosenberg OS, Deindl S, Sung RJ, Nairn AC, Kuriyan J (2005) Structure of the autoinhibited kinase domain of CaMKII and SAXS analysis of the holoenzyme. Cell 123:849-860]. Although the formation of dimers in the intact holoenzyme has not been established, several features of the crystal structure suggest that dimers could act as a synaptic switch. ATP-binding sites were occluded, and the T286 autophosphorylation site responsible for persistent kinase activation was buried. These features would act to stabilize an autoinhibited "paired"-enzyme state. Ca(2+)-calmodulin binding was postulated to trigger the formation of an active state with unpaired catalytic domains. This conformation would allow ATP access and expose T286, autophosphorylation of which would act to maintain the "unpaired" conformation. We used fluorescence anisotropy and FRET imaging of Venus-tagged CaMKIIalpha to test the hypothesis that neuronal CaMKIIalpha can flip between two stable conformations in living cells. Our data support the existence of catalytic domain pairs, and glutamate receptor activation in neurons triggered an increase in anisotropy consistent with a structural transition from a paired to unpaired conformation.
National Academy of Sciences
Journals
2009 EN
Kotaro Miura · Masakazu Agetsuma · Hidemi Kitano
+4 more
Epigenetic mutations confer heritable changes in gene expression that are not due to changes in the underlying sequence of the DNA. We identified a spontaneous rice mutant, Epi-d1, that shows a metastable dwarf phenotype. The phenotype is mitotically and meiotically inheritable and corresponds to the metastable epigenetic silencing of the DWARF1 (D1) gene. The silenced state is correlated with repressive histone and DNA methylation marks in the D1 promoter region but is not associated with DNA sequence alterations. Compared with other known epigenetic silenced loci in plants such as paramutable maize alleles and silent Arabidopsis genes, the Epi-d1 silencing phenomenon shows a high level of bidirectional metastable mutability. Epigenetic alleles such as Epi-d1 could thus provide for rapid adaptation under selective conditions.
National Academy of Sciences
Journals
2009 EN
Andrew D. Redd · Djénéba Dabitao · Jay H. Bream
+16 more
Reports from the United States have demonstrated that elevated markers of microbial translocation from the gut may be found in chronic and advanced HIV-1 infection and are associated with an increase in immune activation. However, this phenomenon's role in HIV-1 disease in Africa is unknown. This study examined the longitudinal relationship between microbial translocation and circulating inflammatory cytokine responses in a cohort of people with varying rates of HIV-1 disease progression in Rakai, Uganda. Multiple markers for microbial translocation (lipopolysaccharide, endotoxin antibody, and sCD14) did not change significantly during HIV-1 disease progression. Moreover, circulating immunoreactive cytokine levels either decreased or remained virtually unchanged throughout disease progression. These data suggest that microbial translocation and its subsequent inflammatory immune response do not have a causal relationship with HIV-1 disease progression in Africa.
National Academy of Sciences
Journals
2009 EN
Jie Ke · WenTa Su · Steven M. Howdle
+13 more
Electrodeposition is a widely used materials-deposition technology with a number of unique features, in particular, the efficient use of starting materials, conformal, and directed coating. The properties of the solvent medium for electrodeposition are critical to the technique's applicability. Supercritical fluids are unique solvents which give a wide range of advantages for chemistry in general, and materials processing in particular. However, a widely applicable approach to electrodeposition from supercritical fluids has not yet been developed. We present here a method that allows electrodeposition of a range of metals from supercritical carbon dioxide, using acetonitrile as a co-solvent and supercritical difluoromethane. This method is based on a careful selection of reagent and supporting electrolyte. There are no obvious barriers preventing this method being applied to deposit a range of materials from many different supercritical fluids. We present the deposition of 3-nm diameter nanowires in mesoporous silica templates using this methodology.
National Academy of Sciences