Further Tests of a Recombination Model in Which χ Removes the Recd Subunit from the Recbcd Enzyme of Escherichia Coli

When one of two infecting lambda phage types in a replication-blocked cross is chi + and DNA packaging is divorced from the RecBCD-chi interaction, complementary chi-stimulated recombinants are recovered equally in mass lysates only if the chi + parent is in excess in the infecting parental mixture. Otherwise, the chi 0 recombinant is recovered in excess. This observation implies that, along with the chi 0 chromosome, two chi + parent chromosomes are involved in the formation of each chi + recombinant. The trimolecular nature of chi +-stimulated recombination is manifest in recombination between lambda and a plasmid. When lambda recombines with a plasmid via the RecBCD pathway, the resulting chromosome has an enhanced probability of undergoing lambda x lambda recombination in the interval into which the plasmid was incorporated. These two observations support a model in which DNA is degraded by Exo V from cos, the sequence that determines the end of packaged lambda DNA and acts as point of entry for RecBCD enzyme, to chi, the DNA sequence that stimulates the RecBCD enzyme to effect recombination. The model supposes that chi acts by ejecting the RecD subunit from the RecBCD enzyme with two consequences. (1) ExoV activity is blocked leaving a highly recombinagenic, frayed duplex end near chi, and (2) as the enzyme stripped of the RecD subunit travels beyond chi it is competent to catalyze reciprocal recombination.     

Identification of glycoinositol phospholipid linked and truncated forms of the scrapie prion protein

Analysis of carboxy-terminal peptides derived from endoproteinase Lys-C digests of the scrapie isoform of the hamster prion protein revealed that the majority of the molecules are glycoinositol phospholipid linked through ethanolamine attached at serin-231. However, approximately 15% of PrPSc had a carboxy-terminal peptide that ends at glycine-228. It is intriguing that this glycine is part of the PrP sequence Gly-Arg-Arg, which is an established target sequence for the proteolysis and release of bioactive peptides from larger precursors. The mechanism of formation, as well as the role of the truncated carboxy terminus in the dissemination and neuropathology of scrapie, remains to be determined.     

Immunocytochemical localization of Na+, K+-ATPase in the rat kidney

To determine if rat kidney Na+, K+-ATPase can be localized by immunoperoxidase staining after fixation and embedding, we prepared rabbit antiserum to purified lamb kidney medulla Na+, K+-ATPase. When sodium dodecylsulfate polyacrylamide electrophoretic gels of purified lamb kidney Na+, K+-ATPase and rat kidney microsomes were treated with antiserum (1:200), followed by [125I]-Protein A and autoradiography, the rat kidney microsomes showed a prominent radioactive band coincident with the alpha-subunit of the purified lamb kidney enzyme and a fainter radioactive band which corresponded to the beta-subunit. When the Na+, K+-ATPase antiserum was used for immunoperoxidase staining of paraffin and plastic sections of rat kidney fixed with Bouin's, glutaraldehyde, or paraformaldehyde, intense immunoreactive staining was present in the distal convoluted tubules, subcapsular collecting tubules, thick ascending limb of the loops of Henle, and papillary collecting ducts. Proximal convoluted tubules stained faintly, and the thin portions of the loops of Henle, straight descending portions of proximal tubules, and outer medullary collecting ducts did not stain. Staining was confined to basolateral surfaces of tubular epithelial cells. No staining was obtained with preimmune serum or primary antiserum absorbed with purified lamb kidney Na+, K+-ATPase, or with osmium tetroxide postfixation. We conclude that the basolateral membranes of the distal convoluted tubules and ascending thick limb of the loops of Henle are the major sites of immunoreactive Na+, K+-ATPase concentration in the rat kidney.     

Chi-Stimulated Recombination between Phage λ and the Plasmid λdv

Chi promotes Rec-mediated recombination between phage lambda DNA and the homologous plasmid lambda dv. In the absence of Chi, some of the interactions splice lambda dv into lambda, whereas others patch information from lambda dv into lambda. When Chi is in the phage DNA, splices and patches are increased in frequency by the same factor. This result strengthens the analogy between Chi and recombination-promoting elements in fungi. It also rules out one model for the previously reported orientation dependence of Chi phenotype.     

Yeast mitochondrial and cytoplasmic valyl-tRNA synthetases

Yeast mitochondrial tRNA synthetase has been partially purified and chromatographic, catalytic and antigenic properties have been compared to the cytoplasmic homologous enzyme from yeast. No significant differences could be observed between the two enzymes with respect to their behaviour during ammonium sulfate precipitation or in chromatographic separation on DEAE cellulose, hydroxylapatite and Sephadex G 200. The Km of the two enzymes toward tRNAs from yeast mitochondria, yeast cytoplasm or E. coli are pratically identical. The antigenic properties of the two enzymes are very similar; antisera against either the mitochondria or the cytoplasmic enzyme lead to the inhibition of their catalytic properties. The mitochondrial ValRS is formed by a single polypeptide chain whose molecular weight is 125,000 daltons, a value very close to that of the yeast cytoplasmic enzyme.     

Clonal growth in vitro by mouse Kupffer cells.

Mouse liver Kupffer cells were induced to proliferate and form discrete colonies of mononuclear phagocytes in vitro. These colony-forming cells from the liver are similar to other mononuclear phagocyte colony-forming cells in that they require a colony-stimulating factor present in medium conditioned by L cells for proliferation in vitro. Cells in the colonies were phagocytic and had IgG receptors on the membrane. For this class of colony-forming cells, the D₀ value to gamma irradiation in vitro was 108 rads.     

Inconsistency in the species concept for yeasts due to mutations during vegetative growth

Summary The generic classification of yeasts is based mainly on morphological characteristics whereas the definition of a species depends predominantly on physiological properties such as the utilization of carbon and nitrogen sources. Classification procedures are routinely done on agar slants, and in negative tests single colonies are often noticed. These colonies are spontaneous mutations and can be idetified as such after transfer onto adequate media and appropriate genetic tests. It is sometimes possible after selection steps to obtain a completely different species. This means that in many cases the classification depends only on single gene differences, where the differences in DNA base homology is almost certainly less than 1%. Since it is rather difficult to justify a new species on the basis of a single biochemical gene mutation, it is necessary in practice to perform at regular intervals an extended series of physiological tests in order to avoid confusion in nomenclature.