Resolution of Dicentric Chromosomes by Ty-Mediated Recombination in Yeast

We have integrated a plasmid containing a yeast centromere, CEN5, into the HIS4 region of chromosome III by transformation. Of the three transformant colonies examined, none contained a dicentric chromosome, but all contained a rearranged chromosome III. In one transformant, rearrangement occurred by homologous recombination between two Ty elements; one on the left arm and the other on the right arm of chromosome III. This event produced a ring chromosome (ring chromosome III) of about 60 kb consisting of CEN3 and all other sequences between the two Ty elements. In addition, a linear chromosome (chromosome IIIA) consisting of sequences distal to the two Ty elements including CEN5, but lacking 60 kb of sequences from the centromeric region, was produced. Two other transformants also contain a similarly altered linear chromosome III as well as an apparently normal copy of chromosome III. These results suggest that dicentric chromosomes cannot be maintained in yeast and that dicentric structures must be resolved for the cell to survive.—The meiotic segregation properties of ring chromosome III and linear chromosome IIIA were examined in diploid cells which also contained a normal chromosome III. Chromosome IIIA and normal chromosome III disjoined normally, indicating that homology or parallel location of the centromeric regions of these chromosomes are not essential for proper meiotic segregation. In contrast, the 60-kb ring chromosome III, which is homologous to the centromeric region of the normal chromosome III, did not appear to pair with fidelity with chromosome III

Meiotic segregation of normal and deletion chromosomes in Saccharomyces cerevisiae.

We explored the behavior of meiotic chromosomes in Saccharomyces cerevisiae by examining the effects of chromosomal rearrangements on recombination and disjunction. Chromosome III derivatives in which the entire left arm or the entire right arm was deleted (telocentric) segregated with fidelity from a normal chromosome III. Recombination between either of these two deletion chromosomes and the intact chromosome also appeared normal. In a strain containing a right arm telocentric, a left arm telocentric and one normal chromosome both telocentrics disjoined from the normal chromosome. Homology on one arm was sufficient for proper recombination and segregation of these chromosomes. In strains containing two normal chromosomes and one telocentric chromosome the two normal chromosomes preferentially disjoined. In a few cases however, the two normal chromosomes cosegregated. Recombination between the two normal chromosomes or between one normal chromosome and the deletion chromosome increased the probability that they would disjoin, although cosegregation of recombinants was observed. A chromosome III derivative which contained a large centromeric deletion and an insertion of the centromere from chromosome V into a nonhomologous position segregated with fidelity from a normal chromosome III. These studies demonstrate that it is not pairing of the centromeres, but pairing and recombination along the arms of the homologs that directs meiotic chromosome disjunction

Treatment of traumatic brain injury using zinc-finger protein gene therapy targeting VEGF-A

Vascular endothelial growth factor (VEGF) plays a role in angiogenesis and has been shown to be neuroprotective following central nervous system trauma. In the present study we evaluated the pro-angiogenic and neuroprotective effects of an engineered zinc-finger protein transcription factor transactivator targeting the vascular endothelial growth factor A (VEGF-ZFP). We used two virus delivery systems, adeno-virus and adeno-associated virus, to examine the effects of early and delayed VEGF-A upregulation after brain trauma, respectively. Male Sprague-Dawley rats were subject to a unilateral fluid percussion injury (FPI) of moderate severity (2.2-2.5atm) followed by intracerebral microinjection of either adenovirus vector (Adv) or an adeno-associated vector (AAV) carrying the VEGF-ZFP construct. Adv-VEGF-ZFP-treated animals had significantly fewer TUNEL positive cells in the injured penumbra of the cortex (p<0.001) and hippocampus (p=0.001) relative to untreated rats at 72h post-injury. Adv-VEGF-ZFP treatment significantly improved fEPSP values (p=0.007) in the CA1 region relative to injury alone. Treatment with AAV2-VEGF-ZFP resulted in improved post-injury microvascular diameter and improved functional recovery on the balance beam and rotarod task at 30 days post-injury. Collectively, the results provide supportive evidence for the concept of acute and delayed treatment following TBI using VEGF-ZFP to induce angiogenesis, reduce cell death, and enhance functional recovery. © Mary Ann Liebert, Inc..link_to_subscribed_fulltex

Characterization of a Recombinant Adeno-Associated Virus Type 2 Reference Standard Material

Here, the AAV Reference Standard Working Group presents the results of their international collaboration to produce and characterize a recombinant AAV serotype 2 Reference Standard Material. The purpose of this material is to provide a universal standard for particle titer, vector genome titer, and infectious titer for AAV2 gene transfer vectors