Cloning and characterization of Hunk, a novel mammalian SNF1-related protein kinase

We previously identified a novel protein kinase, Hunk, by means of a degenerate PCR screen designed to isolate kinases expressed in the murine mammary gland. We now describe the molecular cloning, chromosomal localization, and activity of this kinase and characterize its spatial and temporal pattern of expression in the mouse. We have isolated a 5.0-kb full-length cDNA clone that contains the 714-amino-acid open reading frame encoding Hunk. Analysis of this cDNA reveals that Hunk is most closely related to the SNF1 family of serine/threonine kinases and contains a newly described SNF1 homology domain. Accordingly, antisera specific for Hunk detect an 80-kDa polypeptide with associated phosphotransferase activity. Hunk is located on distal mouse chromosome 16 in a region of conserved synteny with human chromosome 21q22. During fetal development and in the adult mouse, Hunk mRNA expression is developmentally regulated and tissue-specific. Moreover, in situ hybridization analysis reveals that Hunk expression is restricted to subsets of cells within a variety of organs in the adult mouse. These findings suggest a role for Hunk in murine development.     

Reversible sporicidal action of cuprous ions

At 10 mM, Cu⁺ was highly protective against killing of spores of Bacillus megaterium ATCC 19213 by H₂O₂, while at higher concentrations, from 15–100 mM, killing was augmented. In contrast, Cu²⁺, Fe²⁺, Fe³⁺, Co²⁺ or Co³⁺ ions acted only protectively. Cu⁺ itself was sporicidal in the absence of H₂O₂ or ascorbate, and its sporicidal action did not depend on generation of highly reactive oxygen species. It appeared that killing involved either inhibition of germination or copper toxicity to germinated cells in that Cu⁺-inactivated spores did not germinate readily but chemical decoating of the cells prior to plating on a solid medium resulted in reversal of the sporicidal effect. Received 12 July 1996/ Accepted in revised form 03 November 1996     

Selective androgen receptor modulators based on a series of 7H-[1,4]oxazino[3,2-g]quinolin-7-ones with improved in vivo activity

Modification on a lead series of [1,4]oxazino[3,2-g]quinolin-7-ones at the 2-position led to selective androgen receptor modulators with improved in vivo activity. The most potent analog (−)-33a exhibited full maintenance of levator ani muscle at 3 mg/kg and reduced activity on ventral prostate weight in a 2-week orally-dosed and orchidectomized rat maintenance assay.     

Spore heat resistance and specific mineralization

Spores of Bacillus megaterium ATCC 19213, Bacillus subtilis niger and Bacillus stearothermophilus ATCC 7953 were converted to fully demineralized, but viable, H forms by controlled acid titration. H forms were more heat sensitive than were native forms, but z values were greater for killing of H spores than those for native spores. Therefore, the differences in heat sensitivity between native and H forms decreased with increasing killing temperature. The increase in heat sensitivity associated with demineralization did not appear to be due to damage to cortex lytic enzymes of the germination system because it could not be moderated by decoating heated H spores and plating them on medium with added lysozyme. H spores could be remineralized by means of back titration with appropriate base solutions. The remineralized spores, except for the Na form, were then more heat resistant than were H spores. Ca and Mn were more effective in restoring resistance than were Mg and K. Generally, the remineralized forms (except for the Na form) had z values greater than those of the native forms but still less than those of the H forms. At lower killing temperatures, the reinstatement of resistance could be related to the extent of remineralization. However, at higher killing temperatures, only a fraction of the mineral was effective in restoring resistance, and higher levels of remineralization did not result in greater resistance. Mineralization is clearly an important factor in spore heat resistance, but the relationship between resistance and mineralization is complex and dependent on killing temperature.     

Species richness and niche space for temperate and tropical folivores

We measured structural and chemical traits of the leaves of native, broad-leaved trees in two temperate localities [southern Ontario, Canada (34 species), and Missouri (36 species)] and one tropical locality [central Panama (samples of 21 and 23 species)] to test the hypothesis that the greater diversity of tree species and herbivore species in the tropics is associated with greater resource niche space for herbivores. Variables were leaf toughness, water content, dry mass per unit area, several structural and nutritional carbohydrates, common mineral elements, including nitrogen and phosphorus, and several defensive compounds, including tannins and alkaloids. The four samples were almost fully separable by discriminant analysis on the basis of these leaf traits. Variance in log-transformed trait values among species was lowest in the most northern sample, but did not differ significantly between Missouri and Panama. Niche space, estimated as the square root of the total variance in the log-transformed variables within each locality, varied approximately as Panama = 1, Missouri = 0.8, Ontario = 0.5. Although niche space decreases towards higher latitudes, the change does not match the ca. sixfold decrease in tree species richness or the ca. fourfold decrease in Lepidopteran species richness over the latitude range of our samples. Accordingly, tropical folivore diversity is associated with greater resource niche overlap, greater niche specialization, and/or more completely filled niches, or with variation in niche dimensions not measured in this study.     

Enhanced acid resistance of oral streptococci at lethal pH values associated with acid-tolerant catabolism and with ATP synthase activity

Caries-causing oral bacteria such as Streptococcus mutans are protected by the actions of F-ATPases against acid damage in dental plaque acidified by glycolytic acid production or ingestion of acids foods and beverages. Catabolites such as glucose and sucrose were found to enhance the protection of S. mutans and also other oral lactic-acid bacteria against acid killing at lethal pH values as low as 2.5. Protection involved glycolysis with the production of lactate and ATP, which is a substrate for F-ATPases. ATP could also be produced by starved cells apparently through synthase activity of the F-ATPase associated with acid decline. Fluoride and the organic weak-acid indomethacin acted to diminish this protection, as did F-ATPase inhibitors such as dicyclohexylcarbodi-imide. Protection against acid killing involving catabolism and synthase activity is likely to be important for plaque cariogenicity.     

Sporicidal action of peracetic acid and protective effects of transition metal ions

Although peracetic acid (PAA) is used widely for cold sterilization and disinfection, its mechanisms of sporicidal action are poorly understood. PAA at high concentrations (5–10%) can cause major loss of optical absorbance and microscopically-visible damage to bacterial spores. Spores killed by lower levels of PAA (0.02–0.05%) showed no visible damage and remained refractile. Treatment of spores ofBacillus megaterium ATCC 19213 with PAA at concentrations close to the lethal level sensitized the cells to subsequent heat killing. In addition, PAA was found to act in concert with hypochlorite and iodine to kill spores. Antioxidant sulfhydryl compounds or ascorbate protected spores against PAA killing. Trolox, a water-soluble form of -tocopherol, was somewhat protective, while other antioxidants, including -tocopherol, urate, bilirubin, ampicillin and ethanol were not protective. Chelators, including dipicolinate, were not protective, but transition metal ions, especially the reduced forms (Co²⁺, Cu⁺ and Fe²⁺) were highly protective. The net conclusions are that organic radicals formed from PAA are sporicidal and that they may act as reducing agents for spores that are normally in a highly oxidized state, in addition to their well known actions as oxidizing agents in causing damage to vegetative cells.