Evaluation of an FRDA–EGFP genomic reporter assay in transgenic mice

Friedreich ataxia is an autosomal recessive neurodegenerative disorder caused by a GAA trinucleotide expansion in the first intron of the Friedreich ataxia gene (FRDA) that causes reduced synthesis of frataxin, a mitochondrial protein likely to be involved in biosynthesis of iron–sulfur clusters. This leads to increased oxidative stress, progressive loss of large sensory neurons, and hypertrophic cardiomyopathy. To elucidate the mechanisms regulating FRDA expression and to develop an in vivo assay for agents that might upregulate FRDA expression in a therapeutically relevant manner, we have generated transgenic mice with a BAC genomic reporter construct consisting of an in-frame fusion between FRDA and the gene coding for enhanced green fluorescent protein (EGFP). Production of full-length frataxin–EGFP fusion protein was demonstrated by immunoblotting. EGFP expression was observed as early as day E3.5 of development. Most tissues of adult transgenic mice were fluorescent. The level of FRDA–EGFP expression in peripheral blood, bone marrow, and cells obtained from enzymatically disaggregated tissues was quantitated by flow cytometry. There was a twofold increase in EGFP expression in mice homozygous for the transgene when compared to hemizygous mice. These transgenic mice are a valuable tool for the examination of spatial and temporal aspects of FRDA gene expression and for the preclinical evaluation of pharmacological inducers of FRDA expression in a whole-animal model. In addition, tissues from these mice should also be valuable for stem cell transplantation studies.     

A rapid method for determining dynamic binding capacity of resins for the purification of proteins

Biolayer interferometry is a novel method for quantifying macromolecules, such as proteins, in solution. The presence of other, non-binding molecules does not interfere with quantification, which allows one to measure the concentration of the molecule of interest in a crude mixture. Here we apply this method to determining the dynamic binding capacity of affinity resins.     

Developing an approach to defining the potential distributions of invasive plant species: a case study of Hakea species in South Africa

Aim  Models of the potential distributions of invading species have to deal with a number of issues. The key one is the high likelihood that the absence of an invading species in an area is a false absence because it may not have invaded that area yet, or that it may not have been detected. This paper develops an approach for screening pseudo-absences in a way that is logical and defensible.
Innovation  The step-wise approach involves: (1) screening environmental variables to identify those most likely to indicate conditions where the species cannot invade; (2) identifying and selecting the most likely limiting variables; (3) using these to define the limits of its invasion potential; and (4) selecting points outside these limits as true absence records for input into species distribution models.
This approach was adopted and used for the study of three prominent Hakea species in South Africa. Models with and without the false absence records were compared. Two rainfall variables and the mean minimum temperature of the coldest month were the strongest predictors of potential distributions. Models which excluded false absences predicted that more of the potential distribution would have a high invasion potential than those which included them.
Main conclusions  The approach of applying a priori knowledge can be useful in refining the potential distribution of a species by excluding pseudo-absence records which are likely to be due to the species not having invaded an area yet or being undetected. The differences between the potential distributions predicted by the different models convey more information than making a single prediction, albeit a consensus model. The robustness of this approach depends strongly on an adequate knowledge of the ecology, invasion history and current distribution of that species.     

Evidence that Cd101 is an autoimmune diabetes gene in nonobese diabetic mice

We have previously proposed that sequence variation of the CD101 gene between NOD and C57BL/6 mice accounts for the protection from type 1 diabetes (T1D) provided by the insulin-dependent diabetes susceptibility region 10 (Idd10), a <1 Mb region on mouse chromosome 3. In this study, we provide further support for the hypothesis that Cd101 is Idd10 using haplotype and expression analyses of novel Idd10 congenic strains coupled to the development of a CD101 knockout mouse. Susceptibility to T1D was correlated with genotype-dependent CD101 expression on multiple cell subsets, including Foxp3(+) regulatory CD4(+) T cells, CD11c(+) dendritic cells, and Gr1(+) myeloid cells. The correlation of CD101 expression on immune cells from four independent Idd10 haplotypes with the development of T1D supports the identity of Cd101 as Idd10. Because CD101 has been associated with regulatory T and Ag presentation cell functions, our results provide a further link between immune regulation and susceptibility to T1D.     

Trace metals in marine,brackish and freshwater prawns (Crustacea,Decapoda) from northeast Algeria

The concentrations of the trace metals Zn, Cu, Fe, Cd and Pb were measured in four caridean decapods; Atyaephyra desmaresti, Palaemonetes varians, Parapenaeus longirostris and Aristeus antennatus, from freshwater , estuarine and marine habitats in northeast Algeria variably affected by anthropogenic metal contamination. The two coastal species (P. longirostris and A. antennatus) are a food source for the local population. Accumulated metal concentrations varied interspecifically, and intraspecifically between sites and between seasons, except in the case of Aristeus antennatus. The fresh and brackish water species (Atyaephyra desmaresti and Palaemonetes varians) accumulated more Cd and Pb than their marine counterparts. Results are discussed with respect to anthropogenic inputs and the environmental conditions of the regions studied.     

Effects of marginal zinc deficiency on microtubule polymerization in the developing rat brain

One of the possible mechanisms that has been proposed to underlie the deleterious effects of zinc deficiency on brain development is an impairment in the normal formation of the cytoskeletal network. In the current study, in vivo microtubule polymerization was characterized in brain supernatant fluids, from 20-d-old pups whose dams were fed diets containing control (50 micrograms zinc/g) or marginal levels of zinc (10 micrograms zinc/g) throughout pregnancy and lactation. Pup brain and body weights were similar between the groups; however, plasma zinc concentrations were lower (27%) in pups fed the marginal zinc diet than in controls. Tubulin concentrations in 100,000 g brain supernates were similar between the groups; however, tubulin polymerization in the brain supernates was significantly lower in pups fed the marginal zinc diet compared to controls. Primarily, the early events of polymerization were affected; the lag period of the reaction was doubled, and the initial velocity was slower (26%) in supernates from pups fed the marginal zinc diet than in controls. These findings support the idea that some of the negative effects of marginal zinc deficiency on brain development and function may be mediated by an alteration in microtubule formation.