Production of superoxide dismutases from Proteus mirabilis and Proteus vulgaris

Proteus mirabilis and Proteus vulgaris expressed a combination of superoxide dismutase (Sod) activities, which was assigned to FeSod1, FeSod2 and MnSod for P. mirabilis, and FeSod, MnSod and CuZnSod for P. vulgaris. Production of the Sod proteins was dependent on the availability of iron, whether cells were grown under anaerobiosis or aerobiosis and growth phase. Nalidixic acid and chloramphenicol inhibited cell growth and the iron- and dioxygen-dependent production of Sod. These results support the involvement of metal ions and redox status in the production of Proteus Sods.     

Supporting persons with developmental disability--a new model

The way we think about and care for people with developmental disability has changed. Twenty-five years ago, society believed that caregivers always knew what was best for their individual and that he or she must be shielded, even shut away from, the harms that could occur in society. Now, people with disability participate in all aspects of community life. They are educated in local schools, live at home or in their own home, and compete with others in the job market. Caregiving for people with developmental disability is no longer modeled on medicine or stigmatizing labels. Instead, caregivers have become support persons who focus on identifying community resources and making the environment friendlier and safer for persons with disability.     

Introgression and selection shaped the evolutionary history of sympatric sister‐species of coral reef fishes (genus: Haemulon)

Closely related marine species with large overlapping ranges provide opportunities to study mechanisms of speciation, particularly when there is evidence of gene flow between such lineages. Here, we focus on a case of hybridization between the sympatric sister‐species Haemulon maculicauda and H. flaviguttatum, using Sanger sequencing of mitochondrial and nuclear loci, as well as 2422 single nucleotide polymorphisms (SNPs) obtained via restriction site‐associated DNA sequencing (RADSeq). Mitochondrial markers revealed a shared haplotype for COI and low divergence for CytB and CR between the sister‐species. On the other hand, complete lineage sorting was observed at the nuclear loci and most of the SNPs. Under neutral expectations, the smaller effective population size of mtDNA should lead to fixation of mutations faster than nDNA. Thus, these results suggest that hybridization in the recent past (0.174–0.263 Ma) led to introgression of the mtDNA, with little effect on the nuclear genome. Analyses of the SNP data revealed 28 loci potentially under divergent selection between the two species. The combination of mtDNA introgression and limited nuclear DNA introgression provides a mechanism for the evolution of independent lineages despite recurrent hybridization events. This study adds to the growing body of research that exemplifies how genetic divergence can be maintained in the presence of gene flow between closely related species.     

CTCF cis-Regulates Trinucleotide Repeat Instability in an Epigenetic Manner: A Novel Basis for Mutational Hot Spot Determination

At least 25 inherited disorders in humans result from microsatellite repeat expansion. Dramatic variation in repeat instability occurs at different disease loci and between different tissues; however, cis-elements and trans-factors regulating the instability process remain undefined. Genomic fragments from the human spinocerebellar ataxia type 7 (SCA7) locus, containing a highly unstable CAG tract, were previously introduced into mice to localize cis-acting “instability elements,” and revealed that genomic context is required for repeat instability. The critical instability-inducing region contained binding sites for CTCF—a regulatory factor implicated in genomic imprinting, chromatin remodeling, and DNA conformation change. To evaluate the role of CTCF in repeat instability, we derived transgenic mice carrying SCA7 genomic fragments with CTCF binding-site mutations. We found that CTCF binding-site mutation promotes triplet repeat instability both in the germ line and in somatic tissues, and that CpG methylation of CTCF binding sites can further destabilize triplet repeat expansions. As CTCF binding sites are associated with a number of highly unstable repeat loci, our findings suggest a novel basis for demarcation and regulation of mutational hot spots and implicate CTCF in the modulation of genetic repeat instability.     

A High-throughput-compatible FRET-based Platform for Identification and Characterization of Botulinum Neurotoxin Light Chain Modulators

Botulinum neurotoxin (BoNT) is a potent and potentially lethal bacterial toxin that binds to host motor neurons, is internalized into the cell, and cleaves intracellular proteins that are essential for neurotransmitter release. BoNT is comprised of a heavy chain (HC), which mediates host cell binding and internalization, and a light chain (LC), which cleaves intracellular host proteins essential for acetylcholine release. While therapies that inhibit toxin binding/internalization have a small time window of administration, compounds that target intracellular LC activity have a much larger time window of administrations, particularly relevant given the extremely long half-life of the toxin. In recent years, small molecules have been heavily analyzed as potential LC inhibitors based on their increased cellular permeability relative to larger therapeutics (peptides, aptamers, etc.). Lead identification often involves high-throughput screening (HTS), where large libraries of small molecules are screened based on their ability to modulate therapeutic target function. Here we describe a FRET-based assay with a commercial BoNT/A LC substrate and recombinant LC that can be automated for HTS of potential BoNT inhibitors. Moreover, we describe a manual technique that can be used for follow-up secondary screening, or for comparing the potency of several candidate compounds.