Calcium/Calmodulin-Stimulated Protein Kinase II Is Present in Primary Cultures of Cerebral Endothelial Cells

Abstract: Calcium/calmodulin-stimulated protein kinase II (CaMPK II). a major kinase in brain, has been established to play an important role in neurotransmitter release and organization of postsynaptic receptors, and it is known to be involved in long-term potentiation and memory. Less is known about the function of this enzyme in nonneural cells. Here we report on the production, presence, and phosphorylation of the α-subunit of CaM-PK II in primary cultures of cerebral endothelial cells. These results raise the possibility that α-CaM-PK II can act as one of the key enzymes of calcium-mediated intracellular signaling in the cerebral endothelial cells and suggest that α-CaM-PK II may participate in such basic cellular processes as permeability in physiological and pathological conditions.     

The MsK family of alfalfa protein kinase genes encodes homologues of shaggy/glycogen synthase kinase-3 and shows differential expression patterns in plant organs and development

This paper reports on the isolation of a novel class of plant serine/threonine protein kinase genes, MsK-1 , MsK-2 and MsK-3 . They belong to the superfamily of cdc2 -like genes, but show highest identity to the Drosophila shaggy and rat GSK-3 proteins (66–70%). All of these kinases share a highly conserved catalytic protein kinase domain. Different amino-terminal extensions distinguish the different proteins. The different plant kinases do not originate from differential processing of the same gene as is found for shaggy , but are encoded by different members of a gene family. Similarly to the shaggy kinases, the plant kinases show different organ-specific and stage-specific developmental expression patterns. Since the shaggy kinases play an important role in intercellular communication in Drosophila development, the MsK kinases are expected to perform a similar function in plants.     

Culture of and fertile plant regeneration from regenerable embryogenic suspension cell-derived protoplasts of wheat (Triticum aestivum L.)

Summary Regenerable embryogenic cell suspensions initiated from immature embryo-derived friable, fast growing, embryogenic calli of GK Ságvári winter wheat (Triticum aestivum L.) served as sources of protoplasts, which were cultured in different liquid or agarose-solidified media. Protocallus formation was best on KM_8p (Kao and Michayluk 1975) and GM (Li and Murai 1990) media, and protocallus growth on MS (Murashige and Skoog 1962) callus growing medium. Green shoot/plant regeneration occurred on MS regenerating medium, and rooting on MS or N6M (Mórocz et al. 1990) hormone-free media. Protocalli maintained their morphogenic capacity over 4 months, and with multiple subcultures on half-strength MS regenerating medium, the total number of regenerants could be increased. Approximately 1000 shoots/plants were regenerated and over 500 plants were transplanted in the greenhouse. The majority of them had an abnormal chromosome number and low viability, however, one plant grew to maturity and set seed.Abbreviations BAP 6-benzylaminopurine - 2,4-D 2,4-dichlorophenoxyacetic acid - ECS embryogenic cell suspension - GA₃ gibberellic acid - GM General medium - IAA indole-3-acetic acid - IBA indole-3-butyric acid - MS Murashige and Skoog medium - NAA 1-naphthaleneacetic acid - RECS regenerable embryogenic cell suspension     

Molecular characterization and expression of a tobacco histone H1 cDNA

We have isolated a 1104 bp tobacco cDNA clone (H1c12) which includes an 846 bp open reading frame. This encodes a polypeptide of 282 amino acid residues and represents the largest plant H1 histone identified so far. The structure of the deduced protein shows the classical tripartite organization of the H1-type linker histones. The expression of the tobacco H1 histone gene(s) corresponding to the H1c12 cDNA clone was examined during different developmental stages. We found that, at the level of steadystate mRNA, expression of gene(s) encoding this H1 histone was rapidly induced in germinating seeds. The H1 gene was expressed in all tissues examined. However, its expression was higher in tissues known to contain meristematic cells. Furthermore, in the leaves of mature plants accumulation of the H1 mRNA exhibits a very characteristic oscillation. This latter finding indicates that, at least in fully developed plants, the expression of this type of H1 histone gene(s) is modulated by a diurnal cycle.     

Expression of recombinant Renilla luciferase in transgenic plants results in high levels of light emission

A 970 bp DNA fragment which encodes the luciferase enzyme of the marine soft coral Renilla reniformis was fused to the cauliflower mosaic virus (CaMV) promoter. The construct pPCV702-ruc was transferred into alfalfa protoplasts by Ca-PEG-mediated transformation and into tobacco, tomato and potato plants by Agrobacterium -mediated transformation. The light emission from homogenates of alfalfa protoplasts transformed with pPCV702-ruc was 16-fold higher than that of protoplasts transformed with the same vector carrying the bacterial luxF gene. Application of a 3 µM aequous solution of 2-benzyl luciferin (luciferin) on to calli, leaves, roots and slices of tomato fruits and potato tubers of transformed plants resulted in strong light emission within seconds which could be easily visualized by a photon counting camera. Light emissions obtained from tissue homogenates of tobacco plants containing a single copy of the pPCV702-ruc construct were around 20-fold higher than those from plants carrying multiple copies of the firefly luciferase gene and around 360-fold higher than those from plants transformed with the bacterial luciferase gene. Owing to its high efficiency the Renilla luciferase may become a useful and novel tool for gene expression studies in plants and other systems.     

Stable integration of foreign DNA into the chromosome of the cyanobacterium Synechococcus R2

The blue-green alga, Synechococcus R2, is transformed to antibiotic resistance by chimeric DNA molecules consisting of Synechococcus R2 chromosomal DNA linked to antibiotic-resistance genes from Escherichia coli. Chimeric DNA integrates into the Synechococcus R2 chromosome by homologous recombination. The efficiency of transformation, as well as the stability of integrated foreign DNA, depends on the position of the foreign genes relative to Synechococcus R2 DNA in the chimeric molecule. When the Synechococcus R2 DNA fragment is interrupted by foreign DNA, integration occurs through replacement of chromosomal DNA by homologous chimeric DNA containing the foreign insert; transformation is efficient and the foreign gene is stable. Mutagenesis in some cases attends integration, depending on the site of insertion. Foreign DNA linked to the ends of Synechococcus R2 DNA in a circular molecule, however, integrates less efficiently. Integration results in duplicate copies of Synechococcus R2 DNA flanking the foreign gene and the foreign DNA is unstable. Transformation in Synechococcus R2 can be exploited to modify precisely and extensively the genome of this photosynthetic microorganism.