2D/3D hybrid structural model of vocal folds

The spatial dimensionality of the vocal fold vibration is a common challenge in creating parsimonious models of vocal fold vibration. The ideal model is one that is accurate, with the lowest possible computational expense. Inclusion of full 3D flow and structural vibration typically requires massive amounts of computation, whereas reduction of either the flow or the structure to two dimensions eliminates certain aspects of physical reality, thus making the resulting models less accurate. Previous 2D models of the vocal fold structure have utilized a plane strain formulation, which is shown to be an erroneous modeling approach since it ignores influential stress components. We herein present a 2D/3D hybrid vocal fold model that preserves three-dimensional effects of length and longitudinal shear stresses, while taking advantage of a two-dimensional computational domain. The resulting model exhibits static and dynamic responses comparable to a 3D model, and retains the computational advantage of a two-dimensional model.     

Molecular recognition of Candida albicans (1->2)-β-mannan oligosaccharides by a protective monoclonal antibody reveals the immunodominance of internal saccharide residues

A self-consistent model of β-mannan oligosaccharides bound to a monoclonal antibody, C3.1, that protects mice against Candida albicans has been developed through chemical mapping, NMR spectroscopic, and computational studies. This antibody optimally binds di- and trisaccharide epitopes, whereas larger oligomers bind with affinities that markedly decrease with increasing chain length. The (1→2)-β-linked di-, tri-, and tetramannosides bind in helical conformations similar to the solution global minimum. Antibody recognition of the di- and trisaccharide is primarily dependent on the mannose unit at the reducing end, with the hydrophobic face of this sugar being tightly bound. Recognition of a tetrasaccharide involves a frameshift in the ligand interaction, shown by strong binding of the sugar adjacent to the reducing end. We show that frameshifting may also be deliberately induced by chemical modifications. Molecular recognition patterns similar to that of mAb C3.1, determined by saturation transfer difference-NMR, were also observed in polyclonal sera from rabbits immunized with a trisaccharide glycoconjugate. The latter observation points to the importance of internal residues as immunodominant epitopes in (1→2)-β-mannans and to the viability of a glycoconjugate vaccine composed of a minimal length oligosaccharide hapten.     

Molecular basis of dynamic relocalization of Dictyostelium myosin IB

Class I myosins have a single heavy chain comprising an N-terminal motor domain with actin-activated ATPase activity and a C-terminal globular tail with a basic region that binds to acidic phospholipids. These myosins contribute to the formation of actin-rich protrusions such as pseudopodia, but regulation of the dynamic localization to these structures is not understood. Previously, we found that Acanthamoeba myosin IC binds to acidic phospholipids in vitro through a short sequence of basic and hydrophobic amino acids, BH site, based on the charge density of the phospholipids. The tail of Dictyostelium myosin IB (DMIB) also contains a BH site. We now report that the BH site is essential for DMIB binding to the plasma membrane and describe the molecular basis of the dynamic relocalization of DMIB in live cells. Endogenous DMIB is localized uniformly on the plasma membrane of resting cells, at active protrusions and cell-cell contacts of randomly moving cells, and at the front of motile polarized cells. The BH site is required for association of DMIB with the plasma membrane at all stages where it colocalizes with phosphoinositide bisphosphate/phosphoinositide trisphosphate (PIP(2)/PIP(3)). The charge-based specificity of the BH site allows for in vivo specificity of DMIB for PIP(2)/PIP(3) similar to the PH domain-based specificity of other class I myosins. However, DMIB-head is required for relocalization of DMIB to the front of migrating cells. Motor activity is not essential, but the actin binding site in the head is important. Thus, dynamic relocalization of DMIB is determined principally by the local PIP(2)/PIP(3) concentration in the plasma membrane and cytoplasmic F-actin.     

Small leucine rich proteoglycan family regulates multiple signalling pathways in neural development and maintenance

The small leucine-rich repeat proteoglycan (SLRPs) family of proteins currently consists of five classes, based on their structural composition and chromosomal location. As biologically active components of the extracellular matrix (ECM), SLRPs were known to bind to various collagens, having a role in regulating fibril assembly, organization and degradation. More recently, as a function of their diverse proteins cores and glycosaminoglycan side chains, SLRPs have been shown to be able to bind various cell surface receptors, growth factors, cytokines and other ECM components resulting in the ability to influence various cellular functions. Their involvement in several signaling pathways such as Wnt, transforming growth factor-β and epidermal growth factor receptor also highlights their role as matricellular proteins. SLRP family members are expressed during neural development and in adult neural tissues, including ocular tissues. This review focuses on describing SLRP family members involvement in neural development with a brief summary of their role in non-neural ocular tissues and in response to neural injury.     

Altered patterns of fractionation and exon deletions in Brassica rapa support a two-step model of paleohexaploidy

The genome sequence of the paleohexaploid Brassica rapa shows that fractionation is biased among the three subgenomes and that the least fractionated subgenome has approximately twice as many orthologs as its close (and relatively unduplicated) relative Arabidopsis than had either of the other two subgenomes. One evolutionary scenario is that the two subgenomes with heavy gene losses (I and II) were in the same nucleus for a longer period of time than the third subgenome (III) with the fewest gene losses. This "two-step" hypothesis is essentially the same as that proposed previously for the eudicot paleohexaploidy; however, the more recent nature of the B. rapa paleohexaploidy makes this model more testable. We found that subgenome II suffered recent small deletions within exons more frequently than subgenome I, as would be expected if the genes in subgenome I had already been near maximally fractionated before subgenome III was introduced. We observed that some sequences, before these deletions, were flanked by short direct repeats, a unique signature of intrachromosomal illegitimate recombination. We also found, through simulations, that short--single or two-gene--deletions appear to dominate the fractionation patterns in B. rapa. We conclude that the observed patterns of the triplicated regions in the Brassica genome are best explained by a two-step fractionation model. The triplication and subsequent mode of fractionation could influence the potential to generate morphological diversity--a hallmark of the Brassica genus.     

Noncanonical notch signaling modulates cytokine responses of dendritic cells to inflammatory stimuli

Dendritic cell (DC)-derived cytokines play a key role in specifying adaptive immune responses tailored to the type of pathogen encountered and the local tissue environment. However, little is known about how DCs perceive the local environment. We investigated whether endogenous Notch signaling could affect DC responses to pathogenic stimuli. We demonstrate that concurrent Notch and TLR stimulation results in a unique cytokine profile in mouse bone-marrow derived DCs characterized by enhanced IL-10 and IL-2, and reduced IL-12 expression compared with TLR ligation alone. Unexpectedly, modulation of cytokine production occurred through a noncanonical Notch signaling pathway, independent of γ-secretase activity. Modulation required de novo protein synthesis, and PI3K, JNK, and ERK activity were necessary for enhanced IL-2 expression, whereas modulation of IL-10 required only PI3K activity. Further, we show that this γ-secretase-independent Notch pathway can induce PI3K activity. In contrast, expression of the canonical Notch target gene Hes1 was suppressed in DCs stimulated with Notch and TLR ligands simultaneously. Thus, our data suggest that Notch acts as an endogenous signal that modulates cytokine expression of DCs through a noncanonical pathway and therefore has the potential to tailor the subsequent adaptive immune response in a tissue- and/or stage-dependent manner.     

Saliva of Lygus lineolaris digests double stranded ribonucleic acids

The prospects for development of highly specific pesticides based on double stranded ribonucleic acid have been a recent focus of scientific research. Creative applications have been proposed and demonstrated. However, not all insects are sensitive to double stranded RNA (dsRNA) gene knockdown effects; applications in the order Lepidoptera, for example, have met with varied success. Gene knockdown has been demonstrated in several species in the order Hemiptera. In our laboratory, knockdown experiments relied on microinjection of dsRNA into the hemocoel of the tarnished plant bug, Lygus lineolaris. Subsequent experiments delivering dsRNA to insects by feeding were repeatedly unsuccessful in demonstrating knockdown, and a hypothesis was formulated that the dsRNA was digested and degraded by the insect prior to contact with the insect cells. Exposure of dsRNA to insect saliva, insect salivary glands, and insect hemolymph was compared with commercial RNAase III. The saliva of L. lineolaris was found to rapidly digest double stranded RNA. RNAase inhibitor did not affect the activity but heat treatment slowed enzymatic activity.     

Effects of Therapeutic Lifestyle Change diets high and low in dietary fish-derived FAs on lipoprotein metabolism in middle-aged and elderly subjects

The effects of Therapeutic Lifestyle Change (TLC) diets, low and high in dietary fish, on apolipoprotein metabolism were examined. Subjects were provided with a Western diet for 6 weeks, followed by 24 weeks of either of two TLC diets (10/group). Apolipoprotein kinetics were determined in the fed state using stable isotope methods and compartmental modeling at the end of each phase. Only the high-fish diet decreased median triglyceride-rich lipoprotein (TRL) apoB-100 concentration (-23%), production rate (PR, -9%), and direct catabolism (-53%), and increased TRL-to-LDL apoB-100 conversion (+39%) as compared with the baseline diet (all P < 0.05). This diet also decreased TRL apoB-48 concentration (-24%), fractional catabolic rate (FCR, -20%), and PR (-50%) as compared with the baseline diet (all P < 0.05). The high-fish and low-fish diets decreased LDL apoB-100 concentration (-9%, -23%), increased LDL apoB-100 FCR (+44%, +48%), and decreased HDL apoA-I concentration (-15%, -14%) and PR (-11%, -12%) as compared with the baseline diet (all P < 0.05). On the high-fish diet, changes in TRL apoB-100 PR were negatively correlated with changes in plasma eicosapentaenoic and docosahexaenoic acids. In conclusion, the high-fish diet decreased TRL apoB-100 and TRL apoB-48 concentrations chiefly by decreasing their PR. Both diets decreased LDL apoB-100 concentration by increasing LDL apoB-100 FCR and decreased HDL apoA-I concentration by decreasing HDL apoA-I PR.     

RGS16 attenuates pulmonary Th2/Th17 inflammatory responses

The regulators of G protein signaling (RGS) protein superfamily negatively controls G protein-coupled receptor signal transduction pathways. RGS16 is enriched in activated/effector T lymphocytes. In this paper, we show that RGS16 constrains pulmonary inflammation by regulating chemokine-induced T cell trafficking in response to challenge with Schistosoma mansoni. Naive Rgs16(-/-) mice were "primed" for inflammation by accumulation of CCR10(+) T cells in the lung. Upon pathogen exposure, these mice developed more robust granulomatous lung fibrosis than wild-type counterparts. Distinct Th2 or putative Th17 subsets expressing CCR4 or CCR10 accumulated more rapidly in Rgs16(-/-) lungs following challenge and produced proinflammatory cytokines IL-13 and IL-17B. CCR4(+)Rgs16(-/-) Th2 cells migrated excessively to CCL17 and localized aberrantly in challenged lungs. T lymphocytes were partially excluded from lung granulomas in Rgs16(-/-) mice, instead forming peribronchial/perivascular aggregates. Thus, RGS16-mediated confinement of T cells to Schistosome granulomas mitigates widespread cytokine-mediated pulmonary inflammation.     

Rictor regulates phosphorylation of the novel protein kinase C Apl II in Aplysia sensory neurons

J. Neurochem. (2012) 122, 1108-1117. ABSTRACT: Rapamycin-insensitive companion of TOR (Rictor) is a conserved component of target of rapamycin complex 2 (TORC2), a complex implicated in phosphorylation of a number of signal transduction-related kinases, including protein kinase Cs (PKCs) at their 'hydrophobic' site in the carboxy-terminal extension domain. In the marine mollusk, Aplysia californica, an increase in phosphorylation of the novel PKC, Apl II, at the hydrophobic site is associated with a protein synthesis-dependent increase in synaptic strength seen after continuous application of serotonin. To determine if Rictor plays a role in this increase, we cloned the Aplysia ortholog of Rictor (ApRictor). An siRNA-mediated decrease in ApRictor levels in Aplysia sensory neurons led to a decrease in the phosphorylation of PKC Apl II at the hydrophobic site suggesting a role for ApRictor in hydrophobic site phosphorylation. However, over-expression of ApRictor was not sufficient to increase phosphorylation of PKC Apl II. Continuous application of serotonin increased phosphorylation of PKC Apl II at the hydrophobic site in cultured sensory neurons, and this was blocked by Torin, which inhibits both TORC1 and TORC2. Over-expression of ApRictor did not lead to change in the magnitude of serotonin-mediated phosphorylation, but did lead to a small increase in the membrane localization of phosphorylated PKC Apl II. In conclusion, these studies implicate Rictor in phosphorylation of a novel PKC during synaptic plasticity and suggest an additional role for Rictor in regulating the localization of PKCs.