Mutant alpha-synuclein-induced degeneration is reduced by parkin in a fly model of Parkinson's disease.

Parkinson's disease (PD) patients show a characteristic loss of motor control caused by the degeneration of dopaminergic neurons. Mutations in the genes that encode alpha-synuclein and parkin have been linked to inherited forms of this disease. The parkin protein functions as a ubiquitin ligase that targets proteins for degradation. Expression of isoforms of human alpha-synuclein in the Drosophila melanogaster nervous system forms the basis of an excellent genetic model that recapitulates phenotypic and behavioural features of PD. Using this model, we analysed the effect of parkin co-expression on the climbing ability of aging flies, their life span, and their retinal degeneration. We have determined that co-expression of parkin can suppress phenotypes caused by expression of mutant alpha-synuclein. In the developing eye, parkin reduces retinal degeneration. When co-expressed in the dopaminergic neurons, the ability to climb is extended over time. If conserved in humans, we suggest that upregulation of parkin may prove a method of suppression for PD induced by mutant forms of alpha-synuclein.     

The effects of androgens on the β-glands of the tokay (Gekko gecko): Modification of an hypothesis

Study of the posterior abdominal epidermis in hypophysectomized/thyroidectomized male and female tokays following surgery, and subsequent androgen therapy, indicates that, contrary to a previous model, all aspects of β-gland differentiation are under direct androgenic control. On the other hand, another epidermal specialization, the digital foot-pad, shows a pattern of histogenesis directly comparable to that of β-glands, but is unaffected by androgens. These data are discussed with respect to the evolution of glandular epidermal specializations in gekkonid lizards and the possible role of androgens in modifying the control of cell differentiation in lizard epidermis.     

Chromatographic separation of extruded iron-sulfur cores from the apoproteins of Clostridium pasteurianum and spinach ferredoxins in aqueous Triton X-100/urea

Iron-sulfur core extrusions from spinach [( 2Fe-2S]) and Clostridium pasteurianum (2[4Fe-4S]) ferredoxins in aqueous Triton X-100/urea containing excess benzenethiol yield quantitatively [FenSn(SPh)4]2- with n = 2 and n = 4, respectively. The iron-sulfur cluster can be separated from the corresponding apoprotein by rapid passage of the extrusion mixture over a small anaerobic column of Whatman DE-52 anion-exchange cellulose. Essentially quantitative recovery of [FenSn (SPh)4]2- is achieved in the eluate. The apoprotein remaining on the column can be eluted with 0.5 M NaCl. Most of the residual Triton X-100 and benzenethiol can be removed by passage of the apoprotein eluate over a small column of Bio-Beads SM-2, a hydrophobic polystyrene adsorbent. Apoprotein recovery is comparable to that obtained by other chromatographic methods. At least with spinach ferredoxin, the apoprotein prepared in this fashion can be reconstituted. The procedures developed in this work are potentially most applicable to selective removal of [2Fe-2S] and [4Fe-4S] centers from a multicenter enzyme without irreversible denaturation.     

Chromosome transfer and R-prime formation by an RP4::mini-Mu derivative in Escherichia coli, Salmonella typhimurium, Klebsiella pneumoniae, and Proteus mirabilis

We have introduced into the wide host range conjugative plasmid RP4, a mini-Mu derivative which was known to be able to transpose spontaneously in E. coli K-12, and to induce in such a host several kinds of chromosomal rearrangements including replicon fusions. Unlike RP4, RP4::mini-Mu can mediate the transfer of the host chromosome to a recipient bacterium and generate R primes at high frequencies (10^?4 for the transfer of a given marker, 10^?5 for the formation of R primes carrying a given marker). Two such RP4::mini-Mu plasmids were introduced into one Salmonella typhimurium strain, one Klebsiella pneumoniae strain, and one Proteus mirabilis strain. Each of these three strains were mated with an E. coli K-12 recipient and transconjugants carrying R primes were recovered in all three cases at frequencies ranging from 5 × 10^?6 to 10^?7.     

Compartmentation of high-energy phosphates in resting and beating heart cells

The subcellular distribution of ATP, ADP, creatine phosphate and creatine has been analyzed by fast detergent fractionation of isolated frog heart cells. Digitonin fractionation (0.5 mg/ml, 10 s at 2 degrees C in 20 mM 4-morpholinepropanesulfonic acid/3 mM EDTA/230 mM mannitol medium) was used to separate mitochondria and myofilaments from cytosol. To separate myofilaments from the other cellular compartments. Triton X-100 was used (2%, 15 s in the same medium as digitonin). For either resting or beating cells the total cellular contents of ATP, ADP, creatine phosphate and creatine was similar, nevertheless the O2 consumption was 6-times higher. The compartmentation of these metabolites was also identical. Myofilaments contain 1.1 nmol ADP per mg total cellular proteins. In the cytosolic compartment the metabolite concentrations, all measured in nmol per mg total cellular proteins, were: ATP, 13; ADP, 0.25-0.05; creatine phosphate, 18.5 and creatine, 14. This indicated that the reaction catalyzed by creatine kinase was in a state of (or near) equilibrium.     

In vitro anti-HIV activity of phosphorothioate alpha-anomeric oligodeoxynucleotides

Oligonucleotide analogs consisting exclusively of alpha-anomeric deoxynucleoside units bridged with phosphorothioate linkages have been synthesized and tested in vitro as antiviral agents against human immunodeficiency virus (HIV) in human T cells. Two 28-mers, an homopolymer alpha-S-dC28 and an oligomer alpha-S-anti-rev complementary to the initiation site of the regulatory viral gene rev exhibited antiviral activities comparable to those reported for the corresponding beta-anomeric phosphorothioate analogs. In contrast, a nuclease-resistant homopolymer, alpha-dC28 was inactive. Their preliminary results would indicate that the origin of oligonucleotide phosphorothioate anti-HIV activity is not exclusively correlated with their higher nuclease resistance.     

Kinetics of phenylalanine absorption by the rat intestine in vivo after distal resection

The kinetics of L-phenylalanine absorption across rat small intestine in sham and 50% distal resected animals, in vivo, have been studied by perfusing jejunal loops and monitoring the disappearance of the substrate from the perfusate. After 5 months postresection the total phenylalanine absorption was increased. The relationship between total absorption of substrate and its concentration in the bulk phase shows a non-saturable component and a saturable one that can be inhibited by methionine, both in control and remnant jejunum. The slope of the line that represents the non-saturable component is greater in remnant jejunum, indicating that the apparent mass-transfer coefficient, K'D, was increased by distal resection. The kinetic analysis of the saturable component shows that Jmax was unaltered and the apparent semisaturation constant, K'M, was slightly decreased by distal small intestine resection. Correction of the kinetic constant for the unstirred water layer effects shows that the differences between 'real' KD values of the two experimental groups increase whereas 'real' KM values do not change significantly. This indicates that the observed increase in total intestinal absorption in resected animals appears to result from an increase in the intestinal passive permeability.