Hypoxanthine: Guanine phosphoribosyltransferase mutants in Saccharomyces cerevisiae

Summary Yeast mutants lacking activity of the enzyme hypoxanthine: guanine phosphoribosyltransferase (H:GPRT) have been isolated by selecting for resistance to 8-azaguanine in a strain carrying the wild type allele, ade4 ⁺ of the gene coding for amidophosphoribosyltransferase (PRPPAT), the first enzyme of de novo purine synthesis. The mutants excrete purines and are cross-resistant to 8-azaadenine. They are recessive and represent a single complementation group, designated hpt1. Ade4-su, a prototrophic allele of ade4 with reduced activity of PRPPAT, is epistatic to hpt1, suppressing purine excretion and resistance to azaadenine but not resistance to azaguanine. The genotype ade2 hpt1 does not respond to hypoxanthine. Hpt1 complements and is not closely linked to the purine excreting mutants pur1 to pur5. Hpt1 and pur6, a regulatory mutant of PRPPAT, are also unlinked but do not complement, suggesting a protein-protein interaction between H:G-PRT and PRPPAT. Mycophenolic acid (MPA), an inhibitor of de novo guanine nucleotide synthesis, inhibits the growth of hpt1 and hpt1 ⁺. Xanthine allows both genotypes to grow in the presence of MPA whereas guanine only allows growth of hpt1 ⁺. Activity of A-PRT, X-PRT and H:G-PRT is present in hpt ⁺. Hpt1 lacks activity of H:G-PRT but has normal A-PRT and X-PRT.     

Proliferation of a highly androgen-sensitive ductus deferens cell line (DDT1MF-2) is regulated by glucocorticoids and modulated by growth on collagen

Proliferation of the hamster ductus deferens cloned tumor cell line (DDT1MF-2) in monolayer culture is markedly stimulated by androgens in a dose dependent fashion. Furthermore, growth on collagen confers upon these cells a greater dependence on this class of hormones, such that testosterone (10 nM) induces a 15-fold elevation in cell number compared to controls. Addition of either dexamethasone (10 nM) or triamcinolone acetonide (TA; 10 nM) dramatically blocks this stimulation by reversibly arresting the cells in the G1 phase of the cell cycle as assessed by flow cell cytometry. Associated with the decreased growth rate is a change from a rounded to a more flattened morphology that may also implicate cell shape in the regulation of proliferation. These steroid effects presumably are mediated through specific receptor proteins for which dihydrotestosterone (DHT) and TA bind with equilibrium dissociation constants (Kd) of 0.3 and 1.0 nM, respectively. Moreover, not only do androgens increase growth rate but treatment with 1 nM [3H]DHT also results in an elevation in androgen receptor concentration from 1.6 to 3.6 f mol/micrograms DNA in 7 h. Simultaneous treatment with 10 nM TA, however, reduces this increase by 53%. Inasmuch as neither progesterone nor estradiol-17 beta display similar inhibitory activity, this effect also seems to be glucocorticoid specific. These observations may be important in elucidating the mechanism of androgen action and should provide some insight into the role of glucocorticoids in regulating the growth of androgen dependent tissues.     

Restriction map of Tn7

Tn7, a transposon of 14 kb, encodes resistance to trimethoprim (Tp) and streptomycin (Sm). A cleavage site map of this transposon for twenty-two different restriction enzymes as determined by comparison of restriction enzyme cleavage patterns of the plasmids ColE1 and ColE1::Tn7 is presented. The precise localization of these sites was facilitated by the use of two deletion derivatives of ColE1::Tn7: pGB2 and ColE1::Tn7Δ6, and by the use of pOB14 and pOB15 which contain a part of Tn7 cloned into the plasmid pBR322. This map should aid in the study of the structural and genetic organization of this transposon.     

RELATION OF FLOWER COLOR TO OPTICAL-DENSITY SPECTRA OF INTACT TISSUE AND OF ANTHOCYANIN EXTRACTS

Data from measurements of optical density of intact tissue and of anthocyanins in extracts resolved on cellulose thin layer plates were compared with visual evaluations of color quality and intensity in poinsettia, rose, and snapdragon. Visual evaluation was in good agreement with both instrumental and chemical determinations. However, the number or kinds of anthocyanins present could not be predicted from the visual evaluation or from the spectra of the fresh tissue. Data from the resolved extracts did not provide a basis for predicting the optical-density spectrum or the color of the intact tissue. In addition to the genetic factors which have been shown to control (1) the type of anthocyanin, (2) the amount of anthocyanin, and (3) the distribution of anthocyanins within the flower, we suggest another group of genes which apparently affect color through control of structural modification of individual anthocyanins in the living cell through shift in pH, metal chelation, and/or copigmentation. Such genes are apparently responsible for the modification of red color within the Wh Wh genotype of poinsettias containing both pelargonidin and cyanidin glycosides and for a very similar pink color in a snapdragon and a rose, each containing a single anthocyanin, a pelargonidin glycoside, and a cyanidin glycoside, respectively.     

EXPERIMENTAL OBSERVATIONS ON THE INFLUENCE OF TEMPERATURE,LIGHT, AND SALINITY ON CELL DIVISION OF THE MARINE DIATOM,DETONULA CONFERVACEA (CLEVE) GRAN2

The influence of 116 combinations of temperature (2, 7, 12, 16 C), salinity (5–35‰ at 5‰ intervals) and light (5 levels) on the mean daily cell division rate ( K ) of the Narragansett Bay clone of Detonula confervacea was examined following appropriate preconditioning. Growth did not occur at 16 C, but was excellent (K = 1.2–1.5) under certain combinations of light and salinity at 2, 7, and 12 C, being somewhat better at the 2 highest temperature levels. At 32%, and 1100–1200 ft-c, K increased approximately 2.5 fold from 0.6 to 1.5 between 2 and 12 C. A light-temperature relationship was found which had the general trend of an increased optimal light intensity with increasing temperature. Within the optimal salinity range of 15–30‰, the optimal light intensity was 200–600 ft-c at 2 C, 600–1200 ft-c at 7 C, and 1200–1800 ft-c at 12 C. The light-temperature relationship was most pronounced at 2 and 12 C. At 2 C, K decreased with increasing light intensity, but was independent of this factor at higher temperatures. The optimal salinity range of 15–30‰ was independent of temperature negligible growth occurred at 5‰. In situ and in vitro responses of Detonula confervacea to salinity were in general agreement but its pronounced cryophilic preference in nature (usually reaching maximum abundance below 1 C) contrasts with its excellent growth at 12 C in culture. The experiments suggest that termination of the bloom of Detonula confervacea in Narragansett Bay and elsewhere is not solely temperature-dependent. Temperature does not satisfactorily account for its apparent exclusion from waters contiguous to Narragansett Bay and from other more northerly portions of the northeastern coast of the U.S, or, together with light, for its equally surprising apparent unimportance in Norwegian coastal waters.     

Crystalline inclusions in Bacillus thuringiensis

Crystalline inclusion bodies resembling those seen in Clostridium cochlearium were detected in cultures of Bacillus thuringiensis infected with bacteriophage.     

Changes in certain kinetic properties of hepatic microsomal aniline hydroxylase and ethylmorphine demethylase associated with postnatal development and maturation in male rats

1. Changes in certain kinetic properties (V(max.) and apparent K(m)) of hepatic microsomal mixed-function oxidases have been studied as a function of postnatal development and maturation in male rats. 2. Microsomal cytochrome P-450 content changed only slightly between 1 and 12 weeks of age. 3. Aniline hydroxylase activity (V(max.)) increased abruptly between 1 and 2 weeks of age to greater than adult activities and then returned to a plateau value between 4(1/2) and 12 weeks of age. Ethylmorphine demethylase activity remained low and relatively constant between 1 and 3 weeks of age and then increased markedly ( approximately 100%) between 3 and 4(1/2) weeks. 4. The apparent Michaelis constant (K(m)) for aniline hydroxylation increased almost linearly with time between 1 and 6 weeks of age and tended to reach a plateau value thereafter. The apparent K(m) for ethylmorphine demethylation increased between 1 and 3 weeks of age and then decreased abruptly to a constant value between 6 and 12 weeks. 5. The data indicate that developmental changes in the activity of these microsomal oxidases do not correlate temporally with each other or with changes in microsomal cytochrome P-450 content. 6. The most dramatic changes in enzyme activity were associated with early development (1-3 weeks) and weaning (3-4 weeks). 7. Changes in weight of seminal vesicle, a criterion of sexual maturation in male rats, were most prominent between 6 and 8 weeks of age and thus appeared to be separated in time from the prominent changes in enzyme activity.     

Amino acid uptake in the peripheral nerve of the rat

When rat sciatic nerves were incubated with C¹⁴l-lysine, l- or d-glutamate, or d-l γ-aminoisobutyrate, the labeled compounds penetrated the nerve, and the level of lysine and leucine after 1 hr was higher in the nerve than in the medium. The level increased with time, and at 24 hr glutamate levels also were higher in the nerve than in the medium. Lowering the temperature strongly inhibited uptake, while other conditions such as absence of glucose, absence of sodium, or the presence of cyanide inhibited uptake by nerve less than uptake by brain slices. The uptake against a concentration gradient, and inhibitions of this uptake by metabolic inhibitors and by structural analogs, were interpreted as showing the presence of transport processes for amino acids in peripheral nerves with characteristics similar to such transport processes in the central nervous system.