Retrograde Traffic Out of the Yeast Vacuole to the TGN Occurs via the Prevacuolar/Endosomal Compartment

A large number of trafficking steps occur between the last compartment of the Golgi apparatus (TGN) and the vacuole of the yeast Saccharomyces cerevisiae. To date, two intracellular routes from the TGN to the vacuole have been identified. Carboxypeptidase Y (CPY) travels through a prevacuolar/endosomal compartment (PVC), and subsequently on to the vacuole, while alkaline phosphatase (ALP) bypasses this compartment to reach the same organelle. Proteins resident to the TGN achieve their localization despite a continuous flux of traffic by continually being retrieved from the distal PVC by virtue of an aromatic amino acid–containing sorting motif. In this study we report that a hybrid protein based on ALP and containing this retrieval motif reaches the PVC not by following the CPY sorting pathway, but instead by signal-dependent retrograde transport from the vacuole, an organelle previously thought of as a terminal compartment. In addition, we show that a mutation in VAC7, a gene previously identified as being required for vacuolar inheritance, blocks this trafficking step. Finally we show that Vti1p, a v-SNARE required for the delivery of both CPY and ALP to the vacuole, uses retrograde transport out of the vacuole as part of its normal cellular itinerary.     

Calorimetric and Fourier transform infrared spectroscopic studies on the interaction of glycophorin with phosphatidylserine/dipalmitoylphosphatidylcholine-d62 mixtures

Glycophorin has been isolated in pure form from human erythrocyte membranes and reconstituted into lipid vesicles composed of binary mixtures of bovine brain phosphatidylserine (PS) and acyl-chain perdeuterated dipalmitoylphosphatidylcholine (DPPC-d62). The effect of protein on lipid melting behavior and order has been monitored with differential scanning calorimetry and Fourier transform infrared spectroscopy (FT-IR). The phase diagram for PS/DPPC-d62 is consistent with that previously reported for PS/DPPC (Stewart et al. (1979) Biochim. Biophys. Acta 556, 1-16) and indicates that acyl chain perdeuteration does not greatly alter the lipid mixing characteristics. The use of deuterated lipid allows the examination of lipid order by FT-IR of each lipid component in the binary mixtures as well as in the ternary (lipid/lipid/protein) systems. Addition of glycophorin to a 30:70 PS/DPPC-d62 binary lipid mixture results in a preferential glycophorin/PS interaction leading to bulk lipid enriched in DPPC-d62. This is revealed in two ways: first, through cooperative calorimetric transitions increased in temperature from the binary lipid system and second, through FT-IR melting curves of the DPPC-d62 component which shows transitions increased in both onset and completion temperatures in the presence of protein. In addition, non-cooperative melting events are observed at temperatures below the onset of phase separation. The FT-IR data are used to assign these non-cooperative events to the melting of the PS component. For the 50:50 lipid mixture with protein, two transitions are observed in the DSC experiments. The IR results indicate that both lipid components are involved with the lower temperature event.     

Iontophoretic injection of lucifer yellow CH into zygotes and blastomeres of the hydroidHydractinia echinata

The dye Lucifer yellow CH was iontophoresed into recently fertilized eggs and early blastomeres ofHydractinia echinata. Iontophoresis was carried out on the stage of an inverted microscope in order to follow filling of the injected cells by short pulses of epifluorescent illumination. Lucifer yellow proved to be nontoxic and development in embryos with injected blastomeres proceeded normally. When zygotes were injected all the cells of the forming embryo contained dye. When one of the first two blastomeres was injected all the progeny of the injected cell also contained dye. Dye-coupling between injected and uninjected blastomeres did not occur in two-cell embryos nor between descendants of either line. Development of Lucifer-yellow-filled blastomeres or zygotes could be stopped by blue light irradiation. In a number of injected cells, the dye tended to accumulate forming brightly shining spots. The dye did not penetrate the nuclear envelope of injected cells.     

Alkylating Damage by Dipin of Hematopoietic and Stromal Cells of the Bone Marrow

Effect of alkylating agent dipin was studied on hematopoietic (CFU-S) and stromal (CFU-F) progenitor cells. Single administration of dipin (0.06 mg/g) to adult (CBA × C57Bl/6) F1 hybrid mice induced a long-term (2 years) oscillations in the numbers of day 7 CFU-S and day 11 CFU-S in the bone marrow and spleen. Dipin also damaged the hematopoietic stroma as indicated by decreased numbers of CFU-F which remained low for at least a year. The capacity of stromal cells to form ectopic hematopoietic foci was considerably decreased and also remained low for 10 months. The obtained data suggest high dipin sensitivity of the earliest hematopoietic and stromal cells. The dynamics of CFU-S numbers in the hematopoietic organs supports their functioning on the basis of clonal succession (Kay, 1965).__________Translated from Izvestiya Akademii Nauk, Seriya Biologicheskaya, No. 3, 2005, pp. 267–272.Original Russian Text Copyright © 2005 by Domaratskaya, Bueverova, Payushina, Starostin.     

Wives and Warriors: Women and the Military in the United States and Canada

Wives and Warriors: Women and the Military in the United States and Canada. Laurie Weinstein and Christie C. White. eds. Westport, CT: Bergin and Garvey, 1997. 252 pp.     

The effect of microsomal enzyme inducing drugs on reproductive function in the ewe.

A number of drugs cause marked increases in the steroid hydroxylase activity of hepatic microsomes. Beginning 2 days after estrus, 117 mature ewes were each given 14 injections over a 27-day period of phenobarbital sodium, diphenylhydantoin, chlorcyclizine HCl or phenylbutazone. Blood samples for luteinizing hormone (LH) and progesterone determination by radioimmunoassay (RIA) were taken on day 10 of the first estrous cycle (day 18 if no heat was observed) and on days 5 and 10 of the second cycle. On day 10 of the second cycle, the ewes were given an intravenous injection of 1 ml of 6% solution of pentobarbitol sodium anesthetic per 4.5 kg body weight, and the length of anesthetic sleep time was measured. The ewes were then killed and corpora lutea and liver were weighed.In 33 ewes treated with either phenobarbitol sodium or phenylbutazone, sleep time was shortened (18 min $\text{vs}$ 29 min in untreated controls, P<.01), indicating that enzyme induction had occurred. For 41 ewes treated with either chlorcyclizine HCl or diphenylhydantoin, sleep time was lengthened to 93 min (P<.01 $\text{vs}$ controls), indicating impaired liver function. Electron micrographs of liver cells verified that enzyme induction or hepatic degeneration had occurred.