Proliferation-dependent reduction of sister-chromatid exchange frequency induced by mitomycin C in human lymphoblastoid cells and its suppression by inhibitors of DNA replication

A high frequency of sister-chromatid exchange (SCE) induced in cells of a human lymphoblastoid cell line, NL3, by 2-h treatment with 1 microM mitomycin C (MMC) was maintained after holding the treated cells in a nonproliferating state for 48 h before cells were transferred into the BrdUrd-containing medium for SCE assay. The same was observed in cells treated with 4-nitroquinoline 1-oxide (4NQO) or ethyl methanesulfonate (EMS). In contrast, when MMC-treated cells were transferred into a growth medium and allowed to proliferate for various periods of time before SCE assay, MMC-induced SCE frequency decreased with time and reached near control level after 48 h. The reduction in SCE was also observed in 4NQO-treated cells, though to a lesser extent, but not in EMS-treated cells. When hydroxyurea or 1-beta-D-arabinofuranosylcytosine was given as a post-MMC treatment during this recovery process, such a reduction of SCE frequency was suppressed and the extent of the suppression appears to be roughly parallel to their ability to inhibit DNA replication. Cycloheximide and 5-azacytidine also exerted a similar inhibitory effect on the reduction of SCE. Benzamide and caffeine had no appreciable effect. Our results indicate that the SCE-forming lesions induced by MMC can be eliminated only in proliferating cells, probably during DNA replication.     

Three forms of rat liver glucosamine 6-phosphate acetylase and the changes in their levels during development

The three forms (Form I, II and III) of glucosamine 6-phosphate acetylase (glucosamine-phosphate acetyltransferase, EC 2.3.1.4) were present in rat liver. The enzyme activities changed separately during development, in which the successive epigenetic changes were suggested.     

Pneumocystis carinii infection in corticosteroid-treated cats

Corticosteroids were administered to produce Pneumocystis carinii infection in cats. Six of 10 cats, injected intramuscularly for 97-141 days with 2 mg/cat twice weekly of betamethasone sodium phosphate, developed a light infection with P. carinii. Six of 7 cats, injected intramuscularly for 11-168 days with 10-25 mg/cat weekly of prednisolone acetate, also developed a light infection with P. carinii. There was no significant difference in the infection rate between the sexes and ages of the cats. Using Giemsa staining and Gomori's methenamine silver nitrate stain, P. carinii organisms were indistinguishable morphologically from human and rat P. carinii. The cysts and trophozoites were usually present singly or in small groups, and they always were adhering to the periphery of alveoli. The inflammatory changes were inconspicuous except for the fact that alveolar macrophages often were seen. Corticosteroid-treated cats should be useful in the study of experimental P. carinii infection. This is the first reported case of experimentally induced P. carinii infection in cats.     

Cytotoxicity of cysteine in culture media.

When added to Eagle's Minimum Essential Medium supplemented with 10% bovine serum (MEM-10BS), 1mM cysteine was highly toxic to cultured cells. This toxicity was eliminated by (a) preincubation of the medium at 37 degrees C for 24 hr before use, or (b) presence of 5mM pyruvate. Similar results were obtained with freshly prepared CMRL 1066 supplemented with 10% bovine serum (CMRL-10BS), which contains 1.5 mM cysteine as an original ingredient. Medium L 15 supplemented with 10% bovine serum (L-10BS), which contains both 1 mM cysteine and 5 mM pyruvate, supported cell growth. On incubation of MEM-10BS supplemented with 1 mM cysteine (MEM-10BS-1CySH) or CMRL-10BS without cells for one day, the cysteine concentrations decreased to about one-tenth or less of the original concentrations. The cysteine concentration in L-10BS did not decrease so much on similar incubation. Pyruvate reduced the rate of disappearance of the cysteine in MEM-10BS-1CySH or CMRL-10BS as assayed with p-chloromercuribenzoate, although less than that in L-10BS. This effect of pyruvate was concentration dependent. These paradoxical effects of pyruvate on cysteine, i.e. the reduction of its cytotoxicity and the stabilization as an SH compound, are probably due to the formation of a dissociable complex between these two compounds, which is not cytotoxic and resistant to oxidation.     

An endo-(1→6-β-D-glucanase from Mucor hiemalis

An endo-(1→6)-β-D-glucanase (EC 3.2.1), isolated from the culture filtrate of Mucor hiemalis, was purified by ammonium sulphate fractionation and gel filtration. The homogeneity of the enzyme was confirmed by disc electrophoresis. The enzyme had a wide range of temperature and pH stability, high substrate specificity, and an action pattern of the endo-type.     

The plant glycosyltransferase clone collection for functional genomics

The glycosyltransferases (GTs) are an important and functionally diverse family of enzymes involved in glycan and glycoside biosynthesis. Plants have evolved large families of GTs which undertake the array of glycosylation reactions that occur during plant development and growth. Based on the Carbohydrate‐Active enZymes (CAZy) database, the genome of the reference plant Arabidopsis thaliana codes for over 450 GTs, while the rice genome (Oryza sativa) contains over 600 members. Collectively, GTs from these reference plants can be classified into over 40 distinct GT families. Although these enzymes are involved in many important plant specific processes such as cell‐wall and secondary metabolite biosynthesis, few have been functionally characterized. We have sought to develop a plant GTs clone resource that will enable functional genomic approaches to be undertaken by the plant research community. In total, 403 (88%) of CAZy defined Arabidopsis GTs have been cloned, while 96 (15%) of the GTs coded by rice have been cloned. The collection resulted in the update of a number of Arabidopsis GT gene models. The clones represent full‐length coding sequences without termination codons and are Gateway^® compatible. To demonstrate the utility of this JBEI GT Collection, a set of efficient particle bombardment plasmids (pBullet) was also constructed with markers for the endomembrane. The utility of the pBullet collection was demonstrated by localizing all members of the Arabidopsis GT14 family to the Golgi apparatus or the endoplasmic reticulum (ER). Updates to these resources are available at the JBEI GT Collection website http://www.addgene.org/ .