Parasites of Wild Howlers (Alouatta spp.)

A literature review of howler parasites provides the basis for an overview of the ecological significance of parasite surveys in primates. Within this framework, we have added insights into the interactions between primate hosts and their parasites from a long-term study in Costa Rica. We collected fecal samples from mantled howlers (Alouatta palliata) over a 9-year period (1986–1994 inclusive) and analyzed them for parasite eggs, larvae, cysts, and oocysts. We found many misperceptions inherent in the typical methodology of primate parasite surveys and in the reporting of the findings. Our work in Costa Rica suggests that a snapshot effect occurs with most surveys. A static view does not reflect the dynamic and changing ecological interaction between host and parasite. We describe some problems with parasite data analyses that emphasize the need for long-term longitudinal surveys in wild primate groups.     

Bacteria in cancer therapy: a novel experimental strategy

Resistance to conventional anticancer therapies in patients with advanced solid tumors has prompted the need of alternative cancer therapies. Moreover, the success of novel cancer therapies depends on their selectivity for cancer cells with limited toxicity to normal tissues. Several decades after Coley's work a variety of natural and genetically modified non-pathogenic bacterial species are being explored as potential antitumor agents, either to provide direct tumoricidal effects or to deliver tumoricidal molecules. Live, attenuated or genetically modified non-pathogenic bacterial species are capable of multiplying selectively in tumors and inhibiting their growth. Due to their selectivity for tumor tissues, these bacteria and their spores also serve as ideal vectors for delivering therapeutic proteins to tumors. Bacterial toxins too have emerged as promising cancer treatment strategy. The most potential and promising strategy is bacteria based gene-directed enzyme prodrug therapy. Although it has shown successful results in vivo yet further investigation about the targeting mechanisms of the bacteria are required to make it a complete therapeutic approach in cancer treatment.     

Production of haploids and doubled haploids in oil palm

Background

Studies on host-pathogen interactions in a range of pathosystems have revealed an array of mechanisms by which plants reduce the efficiency of pathogenesis. While R-gene mediated resistance confers highly effective defense responses against pathogen invasion, quantitative resistance is associated with intermediate levels of resistance that reduces disease progress. To test the hypothesis that specific loci affect distinct stages of fungal pathogenesis, a set of maize introgression lines was used for mapping and characterization of quantitative trait loci (QTL) conditioning resistance to Setosphaeria turcica, the causal agent of northern leaf blight (NLB). To better understand the nature of quantitative resistance, the identified QTL were further tested for three secondary hypotheses: (1) that disease QTL differ by host developmental stage; (2) that their performance changes across environments; and (3) that they condition broad-spectrum resistance.

Results

Among a set of 82 introgression lines, seven lines were confirmed as more resistant or susceptible than B73. Two NLB QTL were validated in BC₄F₂ segregating populations and advanced introgression lines. These loci, designated qNLB1.02 and qNLB1.06, were investigated in detail by comparing the introgression lines with B73 for a series of macroscopic and microscopic disease components targeting different stages of NLB development. Repeated greenhouse and field trials revealed that qNLB1.06 _Tx303 (the Tx303 allele at bin 1.06) reduces the efficiency of fungal penetration, while qNLB1.02 _B73 (the B73 allele at bin 1.02) enhances the accumulation of callose and phenolics surrounding infection sites, reduces hyphal growth into the vascular bundle and impairs the subsequent necrotrophic colonization in the leaves. The QTL were equally effective in both juvenile and adult plants; qNLB1.06 _Tx303 showed greater effectiveness in the field than in the greenhouse. In addition to NLB resistance, qNLB1.02 _B73 was associated with resistance to Stewart's wilt and common rust, while qNLB1.06 _Tx303 conferred resistance to Stewart's wilt. The non-specific resistance may be attributed to pleiotropy or linkage.

Conclusions

Our research has led to successful identification of two reliably-expressed QTL that can potentially be utilized to protect maize from S. turcica in different environments. This approach to identifying and dissecting quantitative resistance in plants will facilitate the application of quantitative resistance in crop protection.     

Inhibition of protein glycation by skins and seeds of the muscadine grape

The formation of advanced glycation end products (AGEs) leading to protein glycation and cross-linking is associated with the pathogenesis of diabetic complications. The inhibition of protein glycation by phenolic and flavonoid antioxidants demonstrates that the process is mediated, in part, by oxidative processes. In this study, the effects of seed and skin extracts of the muscadine grape on AGEs formation were examined. Seeds and skins were extracted (10% w/v) with 50% ethanol and incubated at 37 degrees C with a solution containing 250 mM fructose and 10 mg/ml albumin. After 72 h, fluorescence was measured at the wavelength pair of 370 and 440 nm as an index of the formation of AGEs. Both seed and skin extracts were found to be efficacious inhibitors of AGE formation. A 1:300 dilution of the seed extract decreased fluorescence by approximately 65%, whereas muscadine grape skin extract produced a 40% lowering. This difference correlates with the greater antioxidant activity found in muscadine seeds in comparison to skins, however, on a mass basis, the inhibitory activities of the seeds and skins were found to be nearly equivalent. Gallic acid, catechin and epicatechin, the three major polyphenols in the seeds, all significantly decreased the AGE product related fluorescence at a concentration of 50 microM. Neither muscadine seed extract nor skin extract inhibited the methylglyoxal-mediated glycation of albumin. These results suggest that consumption of the muscadine grape may have some benefit in altering the progression of diabetic complications.     

Complement component C3d-antigen complexes can either augment or inhibit B lymphocyte activation and humoral immunity in mice depending on the degree of CD21/CD19 complex engagement

C3d can function as a molecular adjuvant by binding CD21 and thereby enhancing B cell activation and humoral immune responses. However, recent studies suggest both positive and negative roles for C3d and the CD19/CD21 signaling complex in regulating humoral immunity. To address whether signaling through the CD19/CD21 complex can negatively regulate B cell function when engaged by physiological ligands, diphtheria toxin (DT)-C3d fusion protein and C3dg-streptavidin (SA) complexes were used to assess the role of CD21 during BCR-induced activation and in vivo immune responses. Immunization of mice with DT-C3d3 significantly reduced DT-specific Ab responses independently of CD21 expression or signaling. By contrast, SA-C3dg tetramers dramatically enhanced anti-SA responses when used at low doses, whereas 10-fold higher doses did not augment immune responses, except in CD21/35-deficient mice. Likewise, SA-C3dg (1 microg/ml) dramatically enhanced BCR-induced intracellular calcium concentration ([Ca2+]i) responses in vitro, but had no effect or inhibited [Ca2+]i responses when used at 10- to 50-fold higher concentrations. SA-C3dg enhancement of BCR-induced [Ca2+]i responses required CD21 and CD19 expression and resulted in significantly enhanced CD19 and Lyn phosphorylation, with enhanced Lyn/CD19 associations. BCR-induced CD22 phosphorylation and Src homology 2 domain-containing protein tyrosine phosphatase-1/CD22 associations were also reduced, suggesting abrogation of negative regulatory signaling. By contrast, CD19/CD21 ligation using higher concentrations of SA-C3dg significantly inhibited BCR-induced [Ca2+]i responses and inhibited CD19, Lyn, CD22, and Syk phosphorylation. Therefore, C3d may enhance or inhibit Ag-specific humoral immune responses through both CD21-dependent and -independent mechanisms depending on the concentration and nature of the Ag-C3d complexes.     

Biosynthetic processing of the Pro-alpha1(V)Pro-alpha2(V)Pro-alpha3(V) procollagen heterotrimer

Type V collagen is a quantitatively minor fibrillar collagen comprised of different chain compositions in different tissues. The most widely distributed form, an alpha1(V)2alpha2(V) heterotrimer, regulates the physical properties of type I/V heterotypic collagen fibrils via partially processed NH2-terminal globular sequences. A less characterized alpha1(V)alpha2(V)alpha3(V) heterotrimer has a much more limited distribution of expression and unknown function(s). We characterized the biosynthetic processing of pro-alpha1(V)2pro-alpha2(V) procollagen previously and showed it to differ in important ways from biosynthetic processing of the major fibrillar procollagens I-III. Here we have successfully produced recombinant pro-alpha1(V)pro-alpha2(V)pro-alpha3(V) heterotrimers. We use these, and mouse embryo fibroblasts doubly homozygous null for the Bmp1 gene, which encodes the metalloproteinase bone morphogenetic protein-1 (BMP-1), and for a gene encoding the closely related metalloproteinase mammalian Tolloid-like 1, to characterize biosynthetic processing of pro-alpha1(V)pro-alpha2(V)pro-alpha3(V) heterotrimers, thus completing characterization of type V collagen biosynthetic processing. Whereas pro-alpha1(V) and pro-alpha2(V) processing in pro-alpha1(V)pro-alpha2(V)pro-alpha3(V) heterotrimers is similar to that which occurs in pro-alpha1(V)2pro-alpha2(V) heterotrimers, the processing of pro-alpha3(V) by BMP-1 occurs at an unexpected site within NH2-terminal globular sequences. We also demonstrate that, despite similarities in NH2-terminal domain structures, pro-alpha2(V) NH2-terminal globular sequences are not cleaved by ADAMTS-2, the metalloproteinase that cleaves the N-propeptides of the major fibrillar procollagen chains.     

BMP1 controls TGFbeta1 activation via cleavage of latent TGFbeta-binding protein

Transforming growth factor beta1 (TGFbeta1), an important regulator of cell behavior, is secreted as a large latent complex (LLC) in which it is bound to its cleaved prodomain (latency-associated peptide [LAP]) and, via LAP, to latent TGFbeta-binding proteins (LTBPs). The latter target LLCs to the extracellular matrix (ECM). Bone morphogenetic protein 1 (BMP1)-like metalloproteinases play key roles in ECM formation, by converting precursors into mature functional proteins, and in morphogenetic patterning, by cleaving the antagonist Chordin to activate BMP2/4. We provide in vitro and in vivo evidence that BMP1 cleaves LTBP1 at two specific sites, thus liberating LLC from ECM and resulting in consequent activation of TGFbeta1 via cleavage of LAP by non-BMP1-like proteinases. In mouse embryo fibroblasts, LAP cleavage is shown to be predominantly matrix metalloproteinase 2 dependent. TGFbeta1 is a potent inducer of ECM formation and of BMP1 expression. Thus, a role for BMP1-like proteinases in TGFbeta1 activation completes a novel fast-forward loop in vertebrate tissue remodeling.     

Mammalian tolloid-like 1 binds procollagen C-proteinase enhancer protein 1 and differs from bone morphogenetic protein 1 in the functional roles of homologous protein domains

Bone morphogenetic protein 1 (BMP1) is the prototype of a subgroup of metalloproteinases with manifold roles in morphogenesis. Four mammalian subgroup members exist, including BMP1 and mammalian Tolloid-like 1 (mTLL1). Subgroup members have a conserved protein domain structure: an NH2-terminal astacin-like protease domain, followed by a fixed order of CUB and epidermal growth factor-like protein-protein interaction motifs. Previous structure/function studies have documented those BMP1 protein domains necessary for secretion, and activity against various substrates. Here we demonstrate that, in contradiction to previous reports, the most NH2-terminal CUB domain (CUB1) is not required for BMP1 secretion nor is the next CUB domain (CUB2) required for enzymatic activity. The same is true for mTLL1. In fact, secreted protease domains of BMP1 and mTLL1, devoid of CUB or epidermal growth factor-like domains, have procollagen C-proteinase (pCP) activity and activity for biosynthetic processing of biglycan, the latter with kinetics superior to those of the full-length proteins. Structure-function analyses herein also suggest differences in the functional roles played by some of the homologous domains in BMP1 and mTLL1. Surprisingly, although BMP1 has long been known to be Ca2+-dependent, a property previously assumed to apply to all members of the subgroup, mTLL1 is demonstrated to be independent of Ca2 levels in its ability to cleave some, but not all, substrates. We also show that pCP activities of only versions of BMP1 and mTLL1 with intact COOH termini are enhanced by the procollagen C-proteinase enhancer 1 (PCOLCE1) and that mTLL1 binds PCOLCE1, thus suggesting reappraisal of the accepted paradigm for how PCOLCE1 enhances pCP activities.     

Modeling haplotype block variation using Markov chains

Models of background variation in genomic regions form the basis of linkage disequilibrium mapping methods. In this work we analyze a background model that groups SNPs into haplotype blocks and represents the dependencies between blocks by a Markov chain. We develop an error measure to compare the performance of this model against the common model that assumes that blocks are independent. By examining data from the International Haplotype Mapping project, we show how the Markov model over haplotype blocks is most accurate when representing blocks in strong linkage disequilibrium. This contrasts with the independent model, which is rendered less accurate by linkage disequilibrium. We provide a theoretical explanation for this surprising property of the Markov model and relate its behavior to allele diversity.