Molecular Mechanisms of Disease-Causing Missense Mutations

Genetic variations resulting in a change of amino acid sequence can have a dramatic effect on stability, hydrogen bond network, conformational dynamics, activity and many other physiologically important properties of proteins. The substitutions of only one residue in a protein sequence, so-called missense mutations, can be related to many pathological conditions and may influence susceptibility to disease and drug treatment. The plausible effects of missense mutations range from affecting the macromolecular stability to perturbing macromolecular interactions and cellular localization. Here we review the individual cases and genome-wide studies that illustrate the association between missense mutations and diseases. In addition, we emphasize that the molecular mechanisms of effects of mutations should be revealed in order to understand the disease origin. Finally, we report the current state-of-the-art methodologies that predict the effects of mutations on protein stability, the hydrogen bond network, pH dependence, conformational dynamics and protein function.     

Nematode vectors of plant viruses in New Zealand

Abstract

A survey of virus vector nematodes based on a total of 320 published and recent records revealed that 3 Longidorus, 1 Paralongidorus, 6 Xiphinema, 2 Trichodoms, and 3 Paratrichodorus species are present in New Zealand. Xiphinema diversicaudatum and Paratrichodorus minor are the most common and widespread species. Longidorus orongorongensis, an undescribed Paralongidorus species and Trichodoms cottieri appear to be endemic species, and Longidorus taniwha, Paratrichodorus lobatus and probably one (still unidentified) species of the X. americanum group are also indigenous to New Zealand. Most of the other species (e.g. Longidorus elongatus, X. diversicaudatum, and Trichodoms primitivus) were probably introduced from Europe. Maps showing the distribution of the 15 species in New Zealand, and illustrations and tables of the morphological characters for species identification, are presented. Several species are of economic importance through direct damage they cause to cultivated plants, and all can be considered potential vectors of plant viruses.     

Insertion of Unmarked DNA Sequences in Multiple Loci of the Bacillus subtilis 168 Genome: an Efficient Selection Method

The construction process of Bacillus subtilis strain with multiple mutations is accelerated by simultaneous use of a positive selection method. All the strains selected by neomycin acquired two simultaneous mutations at different genomic loci. The extremely low rate of false positives was accounted for by employing the improved method.     

A model for Chagas disease with controlled spraying

Chagas disease is a vector-borne parasitic disease that infects mammals, including humans, through much of Latin America. This work presents a mathematical model for the dynamics of domestic transmission in the form of four coupled nonlinear differential equations. The four equations model the number of domiciliary vectors, the number of infected domiciliary vectors, the number of infected humans, and the number of infected domestic animals. The main interest of this work lies in its study of the effects of insecticide spraying and of the recovery of vector populations with cessation of spraying. A novel aspect in the model is that yearly spraying, which is currently used to prevent transmission, is taken into account. The model's predictions for a representative village are discussed. In particular, the model predicts that if pesticide use is discontinued, the vector population and the disease can return to their pre-spraying levels in approximately 5–8 years.     

Adenovirus-mediated delivery of human IFNgamma gene inhibits prostate cancer growth

Interferon gamma (IFNgamma) is regarded as a potent antitumor agent, but therapy with IFNgamma is hampered by its short half-life and significant side effects. We developed a replication defective adenovirus carrying the human IFNgamma gene and evaluated the effects of adenovirus-mediated IFNgamma (Ad-IFNgamma) gene transfer on human prostate cancer cell lines in vitro and on xenografts in vivo. Our results showed infection of prostate cancer cells with Ad-IFNgamma led to production of an active cytokine and resulted in an antiproliferative effect on the prostate cancer cells. Intratumoral injection of Ad-IFNgamma significantly inhibited the growth of DU-145 cell xenografts in vivo, while no significant toxicity effect was observed. RT-PCR analysis indicated transgene expression mainly enriched in tumors in vivo, and slightly distributed in livers. These findings suggest adenovirus-mediated IFNgamma gene transfer is a promising approach in the treatment of advanced prostate cancer.     

Bvui: a site-specific endonuclease from Bacillus vulgatis

A site-specific endodeoxyribonuclease was partially purified from an extract of osmotically shocked spheroplasts of a Bacillus vulgatis strain; the enzyme has been designated BvuI and its activity was characterized. The locations of BvuI-generated cleavages on bacteriophage lambda and M13 derivatives mp7, mp8 and mp9, SV40 and PBR322 DNA molecules were determined. BvuI was shown to recognize the DNA sequence decreases 5'-G-Pu-G-C-Py-C-3' 3'-C-Py-C-G-Pu-G-5' increases and cleaves it at the positions indicated by arrows. Two identical BvuI recognition sites separated by fourteen nucleotide pairs were shown to occur within the tetracycline resistance gene of PBR322; BvuI should be useful for molecular cloning in that plasmid vector.     

Transformation of Aspergillus nidulans using the argB gene

Transformation of Aspergillus nidulans has been achieved using a chimeric vector comprising Escherichia coli, Saccharomyces cerevisiae and Aspergillus nidulans DNA. Protoplasts of argB^? strains (defective for the ornithine carbamoyl transferase [carbamoylphosphate: l-ornithine carbamoyltransferase, EC 2.1.3.3] gene) of A. nidulans were incubated with plasmid pSal43 containing the cloned argB⁺ gene in the presence of poly(ethylene glycol) and CaCl₂. Transformant progeny was of three types; the majority were small slow-growing colonies which were non-viable when transferred to MM. The remaining large colonies, which were recovered at a frequency of 50 μg^?1 DNA in the best experiments, made up the other two types. One group were mitotically stable, showing no evidence of instability; the other comprised unstable types which segregated apparent transformant and parental phenotypes. The apparent transformants showed similar segregational properties. Southern hybridizations with a stable transformant suggested that it arose following integration of the argB⁺ at the arg locus. Analysis of an unstable transformant suggested that possibly more than one copy of the plasmid was integrated and then subjected to rearrangement.     

Vector competence of Coquillettidia linealis (Skuse) (Diptera: Culicidae) for Ross River and Barmah Forest viruses

Abstract Coquillettidia linealis is a severe pest on some of the Moreton Bay islands in Queensland, Australia, but little is known of its breeding habitats and biology. Because of its high abundance and its association with Ross River (RR) and Barmah Forest (BF) viruses by field isolation, its vector competence was evaluated in the laboratory by feeding dilutions of both viruses in blood. For RR, Cq. linealis was of comparable efficiency to Ochlerotatus vigilax (Skuse), recognised as being a major vector. Results were as follows for Cq. linealis and Oc. vigilax , respectively: dose to infect 50%, 10^2.2 and <10^1.7 CCID₅₀/mosquito; 88% and 90% disseminated infection at 4 days postinfection; transmission at 4 days with rates of 68−92% and 25−60%. For BF dose to infect 50%, 10^2.7 and 10^2.0; disseminated infection rates on first transmission day (day 6), 40% and 70%; transmission rates of 8−16% and 0−10%. As a capillary-tube method was used rather than suckling mice to demonstrate transmission, transmission rates may be underestimates. This, the first study of the vector competence of Cq. linealis in Australia, demonstrates that this species deserves control on the southern Moreton Bay islands.     

Gene therapy with the TRF1 telomere gene rescues decreased TRF1 levels with aging and prolongs mouse health span

The shelterin complex protects telomeres by preventing them from being degraded and recognized as double‐strand DNA breaks. TRF1 is an essential component of shelterin, with important roles in telomere protection and telomere replication. We previously showed that TRF1 deficiency in the context of different mouse tissues leads to loss of tissue homeostasis owing to impaired stem cell function. Here, we show that TRF1 levels decrease during organismal aging both in mice and in humans. We further show that increasing TRF1 expression in both adult (1‐year‐old) and old (2‐year‐old) mice using gene therapy can delay age‐associated pathologies. To this end, we used the nonintegrative adeno‐associated serotype 9 vector (AAV9), which transduces the majority of mouse tissues allowing for moderate and transient TRF1 overexpression. AAV9‐TRF1 gene therapy significantly prevented age‐related decline in neuromuscular function, glucose tolerance, cognitive function, maintenance of subcutaneous fat, and chronic anemia. Interestingly, although AAV9‐TRF1 treatment did not significantly affect median telomere length, we found a lower abundance of short telomeres and of telomere‐associated DNA damage in some tissues. Together, these findings suggest that rescuing naturally decreased TRF1 levels during mouse aging using AAV9‐TRF1 gene therapy results in an improved mouse health span.     

基于纳米基因载体的动植物遗传转化研究进展

转基因技术在动植物优良新品种的培育中发挥着重要作用,而随着纳米生物技术的发展,基于纳米材料构建基因载体的动植物转基因技术,对于发展动植物转基因新方法以及加速转基因种质材料的大规模制备、优良新品种的培育进程具有更为重要的意义。综述了纳米基因载体的种类与性质,并结合动植物遗传育种的研究进展,分析了纳米基因载体相比于其他载体的特点及优势,同时,重点阐述了基于纳米基因载体的基因转染技术的基本原理和操作过程,及其在动植物遗传转化中的应用,以期为动植物基因工程改造提供新思路。