Altered Mitochondrial Respiration in Selectively Vulnerable Brain Subregions Following Transient Forebrain Ischemia in the Rat

Mitochondrial respiratory function, assessed from the rate of oxygen uptake by homogenates of rat brain subregions, was examined after 30 min of forebrain ischemia and at recirculation periods of up to 48 h. Ischemia-sensitive regions which develop extensive neuronal loss during the recirculation period (dorsal-lateral striatum, CA1 hippocampus) were compared with ischemia-resistant areas (paramedian neocortex, CA3 plus CA4 hippocampus). All areas showed reductions (to 53-69% of control) during ischemia for oxygen uptake rates determined in the presence of ADP or an uncoupling agent, which then recovered within 1 h of cerebral recirculation. In the ischemia-resistant regions, oxygen uptake rates remained similar to control values for at least 48 h of recirculation. After 3 h of recirculation, a significant decrease in respiratory activity (measured in the presence of ADP or uncoupling agent) was observed in the dorsal-lateral striatum which progressed to reductions of greater than 65% of the initial activity by 24 h. In the CA1 hippocampus, oxygen uptake rates were unchanged for 24 h, but were significantly reduced (by 30% in the presence of uncoupling agent) at 48 h. These alterations parallel the development of histological evidence of ischemic cell change determined previously and apparently precede the appearance of differential changes between sensitive and resistant regions in the content of high-energy phosphate compounds. These results suggest that alterations of mitochondrial activity are a relatively early change in the development of ischemic cell death and provide a sensitive biochemical marker for this process.     

Engineered biosynthesis of bacteriochlorophyll gF in Rhodobacter sphaeroides

Engineering photosynthetic bacteria to utilize a heterologous reaction center that contains a different (bacterio) chlorophyll could improve solar energy conversion efficiency by allowing cells to absorb a broader range of the solar spectrum. One promising candidate is the homodimeric type I reaction center from Heliobacterium modesticaldum. It is the simplest known reaction center and uses bacteriochlorophyll (BChl) g, which absorbs in the near-infrared region of the spectrum. Like the more common BChls a and b, BChl g is a true bacteriochlorin. It carries characteristic C3-vinyl and C8-ethylidene groups, the latter shared with BChl b. The purple phototrophic bacterium Rhodobacter (Rba.) sphaeroides was chosen as the platform into which the engineered production of BChl g_F, where F is farnesyl, was attempted. Using a strain of Rba. sphaeroides that produces BChl b_P, where P is phytyl, rather than the native BChl a_P, we deleted bchF, a gene that encodes an enzyme responsible for the hydration of the C3-vinyl group of a precursor of BChls. This led to the production of BChl g_P. Next, the crtE gene was deleted, thereby producing BChl g carrying a THF (tetrahydrofarnesol) moiety. Additionally, the bchG^Rs gene from Rba. sphaeroides was replaced with bchG^Hm from Hba. modesticaldum. To prevent reduction of the tail, bchP was deleted, which yielded BChl g_F. The construction of a strain producing BChl g_F validates the biosynthetic pathway established for its synthesis and satisfies a precondition for assembling the simplest reaction center in a heterologous organism, namely the biosynthesis of its native pigment, BChl g_F.     

DNA topoisomerase II, genotoxicity, and cancer

Type II topoisomerases are ubiquitous enzymes that play essential roles in a number of fundamental DNA processes. They regulate DNA under- and overwinding, and resolve knots and tangles in the genetic material by passing an intact double helix through a transient double-stranded break that they generate in a separate segment of DNA. Because type II topoisomerases generate DNA strand breaks as a requisite intermediate in their catalytic cycle, they have the potential to fragment the genome every time they function. Thus, while these enzymes are essential to the survival of proliferating cells, they also have significant genotoxic effects. This latter aspect of type II topoisomerase has been exploited for the development of several classes of anticancer drugs that are widely employed for the clinical treatment of human malignancies. However, considerable evidence indicates that these enzymes also trigger specific leukemic chromosomal translocations. In light of the impact, both positive and negative, of type II topoisomerases on human cells, it is important to understand how these enzymes function and how their actions can destabilize the genome. This article discusses both aspects of human type II topoisomerases.     

Genome-wide meta-analysis identifies regions on 7p21 (AHR) and 15q24 (CYP1A2) as determinants of habitual caffeine consumption

We report the first genome-wide association study of habitual caffeine intake. We included 47,341 individuals of European descent based on five population-based studies within the United States. In a meta-analysis adjusted for age, sex, smoking, and eigenvectors of population variation, two loci achieved genome-wide significance: 7p21 (P = 2.4 × 10(-19)), near AHR, and 15q24 (P = 5.2 × 10(-14)), between CYP1A1 and CYP1A2. Both the AHR and CYP1A2 genes are biologically plausible candidates as CYP1A2 metabolizes caffeine and AHR regulates CYP1A2.     

Tandem quadruplication of HMA4 in the zinc (Zn) and cadmium (Cd) hyperaccumulator Noccaea caerulescens

Zinc (Zn) and cadmium (Cd) hyperaccumulation may have evolved twice in the Brassicaceae, in Arabidopsis halleri and in the Noccaea genus. Tandem gene duplication and deregulated expression of the Zn transporter, HMA4, has previously been linked to Zn/Cd hyperaccumulation in A. halleri. Here, we tested the hypothesis that tandem duplication and deregulation of HMA4 expression also occurs in Noccaea.A Noccaea caerulescens genomic library was generated, containing 36,864 fosmid pCC1FOS™ clones with insert sizes ∼20–40 kbp, and screened with a PCR-generated HMA4 genomic probe. Gene copy number within the genome was estimated through DNA fingerprinting and pooled fosmid pyrosequencing. Gene copy numbers within individual clones was determined by PCR analyses with novel locus specific primers. Entire fosmids were then sequenced individually and reads equivalent to 20-fold coverage were assembled to generate complete whole contigs.Four tandem HMA4 repeats were identified in a contiguous sequence of 101,480 bp based on sequence overlap identities. These were flanked by regions syntenous with up and downstream regions of AtHMA4 in Arabidopsis thaliana. Promoter-reporter β-glucuronidase (GUS) fusion analysis of a NcHMA4 in A. thaliana revealed deregulated expression in roots and shoots, analogous to AhHMA4 promoters, but distinct from AtHMA4 expression which localised to the root vascular tissue.This remarkable consistency in tandem duplication and deregulated expression of metal transport genes between N. caerulescens and A. halleri, which last shared a common ancestor >40 mya, provides intriguing evidence that parallel evolutionary pathways may underlie Zn/Cd hyperaccumulation in Brassicaceae.     

The matrilineal ancestry of Ashkenazi Jewry: portrait of a recent founder event

Both the extent and location of the maternal ancestral deme from which the Ashkenazi Jewry arose remain obscure. Here, using complete sequences of the maternally inherited mitochondrial DNA (mtDNA), we show that close to one-half of Ashkenazi Jews, estimated at 8,000,000 people, can be traced back to only 4 women carrying distinct mtDNAs that are virtually absent in other populations, with the important exception of low frequencies among non-Ashkenazi Jews. We conclude that four founding mtDNAs, likely of Near Eastern ancestry, underwent major expansion(s) in Europe within the past millennium.     

The Kinetochore-Microtubule Coupling Machinery Is Repurposed in Sensory Nervous System Morphogenesis

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