Nucleosome-associated protein kinases in murine erythroleukemia cells.

Chromatin subunits from murine erythroleukemia cells were prepared by a method which releases actively transcribing genes. Two casein kinase activities (CK₁ and CK₂) were isolated from these nucleosomes by gel nitration in 0.5 m NaCl. CK₁ (M_r ~ 200,000) and CK₂ (M_r ~ 35,000) were further purified by phosphocellulose chromatography and characterized with regard to several parameters which may regulate their activity in vivo. CK₁ has an NaCl optimum of 0.14 m, utilizes GTP as phosphate donor ~25% as efficiently as ATP, and phosphorylates a discrete group of high molecular weight nonhistone proteins in the unfractionated chromatin starting material. CK₂ has an NaCl optimum of 0.24 m, cannot utilize GTP, and modifies a different group of nonhistones. Both kinases are inhibited by concentrations of hemin (<50 μm) which efficiently induce globin gene expression in erythroleukemia cells. A histone kinase resolved during the gel filtration step is unaffected by hemin. An investigation of the mode of hemin inhibition reveals that CK₁ and CK₂ interact in different fashions with the inhibitor.     

Sugarbeet leaf disc culture: an improved procedure for inducing morphogenesis

In preparation for gene transfer experiments we investigated factors that might affect the production of shoots and somatic embryos from the wound callus of cultured sugarbeet leaf discs. A complex interaction was found between the leaf disc plating density, the disc culture medium, the source-shoot culture medium and the frequency of disc transfer to fresh medium. The most productive protocol utilized: source shoots maintained on MS medium containing 0.25 mg 1^-1 BA; multiple leaf discs (ten 4-mm discs/plate) plated onto an enriched modification of MS medium (RV) containing 1.0 mg 1^-1 BA and solidified with 0.3% Gelrite (not permitted to dry during hardening); and transfer of the discs to fresh medium every two weeks during the first month. This standard protocol produced more callus per plate and higher rates of morphogenesis per unit dry weight of callus than did the one-step method of Saunders and Doley. Water availability considerations were found to be critical to obtaining high morphogenic rates. Root induction frequency and quality was superior on shoots transplanted to MS medium containing 1 mg 1^-1 NAA as the sole growth regulator compared to IAA at the same concentration.Abbreviations BA N⁶-benzyladenine - IAA indole-3-acetic acid - NAA -naphthaleneacetic acid     

Mouse pulmonary cytochrome P-450 naphthalene hydroxylase: cDNA cloning, sequence, and expression in Saccharomyces cerevisiae.

We have isolated a cDNA clone, Nah-2, encoding the cytochrome P-450Nah (naphthalene hydroxylase) from a mouse lung lambda ZAP cDNA library using anti-cytochrome P-450Nah IgG as a probe. This same antibody selectively blocked [Nagata, K., Martin, B.M., Gillette, J.R., & Sasame, H.A. (1990) Drug Metab. Dispos. 18, 557-564] the cytochrome P-450 in mouse lung microsomes that catalyzed the conversion of naphthalene to (1R,2S)-naphthalene 1,2-oxide, which has been postulated as a causative agent in the naphthalene-induced tissue-specific necrosis of Clara cells in mouse lung. The toxic effect is seen in mouse and not in rat. The cDNA encodes a polypeptide of 491 amino acids with a molecular mass of 50 kDa. Northern blot analysis with an Nah-2-specific probe revealed that the mRNA is expressed in a species- and tissue-specific manner, present only in mouse lung and liver and not in that of rat. The mRNA encoding Nah-2 is constitutively expressed and is not induced by either phenobarbital, pyrazole, pregnenolone 16 alpha-carbonitrile, or 3-methylcholanthrene. Comparative amino acid sequence analyses with other documented members of the P-450 gene superfamily revealed that this encoded protein is in the IIF subfamily. To analyze its substrate specificity, the cDNA was inserted into the vector, pAAH5, and expressed in the Saccharomyces cerevisiae strain, AH22. The presence of cytochrome P-450Nah in the microsomes isolated from transformed cells and analyzed by Western blot was confirmed by immunocomplexing product with anti-cytochrome P450Nah IgG. Furthermore, activity toward naphthalene in the microsomes from the transformed cells established that this clone encodes a naphthalene hydroxylase.(ABSTRACT TRUNCATED AT 250 WORDS)     

Characteristics of teratomas regenerated in vitro from octopine-type crown gall

Crown galls induced by infection of tobacco plants with Agrobacterium tumefaciens strain C58-Cl(pTiB6S3) were excised and cultured in vitro. After about one year of culture on medium-lacking phytohormones, two noncloned lines spontaneously formed shoots. Leaf explants from shoots of tumor-line T5 were capable of growing on hormone-free medium, and the resulting mixture of organized and unorganized tissue synthesized octopine. Detached leaves from T5 shoots also synthesized octopine. These results establish that shoots from this octopine-type tumor contain transformed cells and are true crown-gall teratomas.     

Evolutionary ecology of opsin gene sequence,expression and repertoire

Linking molecular evolution to biological function is a long‐standing challenge in evolutionary biology. Some of the best examples of this involve opsins, the genes that encode the molecular basis of light reception. In this issue of Molecular Ecology, three studies examine opsin gene sequence, expression and repertoire to determine how natural selection has shaped the visual system. First, Escobar‐Camacho et al. ( 2017 ) use opsin repertoire and expression in three Amazonian cichlid species to show that a shift in sensitivity towards longer wavelengths is coincident with the long‐wavelength‐dominated Amazon basin. Second, Stieb et al. ( 2017 ) explore opsin sequence and expression in reef‐dwelling damselfish and find that UV‐ and long‐wavelength vision are both important, but likely for different ecological functions. Lastly, Suvorov et al. ( 2017 ) study an expansive opsin repertoire in the insect order Odonata and find evidence that copy number expansion is consistent with the permanent heterozygote model of gene duplication. Together these studies emphasize the utility of opsin genes for studying both the local adaptation of sensory systems and, more generally, gene family evolution.