L J Van Eldik

Biosynthesis of calmodulin in normal and virus-transformed chicken embryo fibroblasts.

We report here that the higher levels of calmodulin in transformed chicken embryo fibroblasts are due to an increase in the rate of synthesis of calmodulin that results from an increased amount of calmodulin-specific mRNA in transformed cells. Transformation of several types of eucaryotic cells by oncogenic viruses results in a two- to threefold increase in the intracellular levels of calmodulin. We used the normal chicken embryo fibroblast and its Rous sarcoma virus-transformed counterpart to examine the biosynthesis of calmodulin. We show that the higher levels of calmodulin found in transformed fibroblasts appear to be the consequence of a selective increase in the rate of synthesis of calmodulin above that of total soluble or total cellular protein. A significant difference in the rate of degradation of calmodulin or total protein between transformed and normal cells was not detected. We also examined the mechanism of the increased synthesis rate of calmodulin and show that the levels of calmodulin mRNA are increased in transformed fibroblasts as measured by both translational activity and hybridization to a calmodulin cDNA probe. It is suggested by these data that the higher levels of calmodulin in transformed cells may result from a specific increase in the rate of either calmodulin gene transcription or mRNA processing.

Perturbation of the calmodulin system in transformed cells.

In eukaryotic cells, calcium acts as an intracellular signal transducer primarily through its interaction with a class of calcium binding proteins, of which calmodulin (CaM) is the most highly conserved, phylogenetically ubiquitous member (for reviews, see Van Eldik et al., 1982; Van Eldik and Roberts, 1988; Cohen and Klee, 1988). CaM transduces a calcium signal into a biological response by its ability to regulate the activity of other proteins in a calcium dependent manner. Because CaM is probably the most widely distributed mediator of intracellular calcium signals, fundamental insights can be derived from an enhanced knowlege about the genetic encoding, biosynthetic assembly and regulation of CaM-modulated calcium response pathways. Regardless, in order to understand fully the roles of CaM in the eukaryotic cell and obtain insight into how CaM-modulated pathways can respond differentially to calcium signals, it is necessary to be able to describe in some detail all of the CaM pathways for at least one biological system. This has not been done yet for any biological system. Chicken embryo fibroblasts (CEF) represent one biological system for which a relatively extensive body of information has been described. Also, CEF transformed by Rous sarcoma virus exhibit a number of phenotypic alterations that are potentially mediated by CaM and CaM binding proteins, and perturbations in CaM regulation have been described for normal and transformed CEF. In a more general sense, because perturbations of CaM pathways occur in many kinds of virus-transformed cells, knowledge of how alterations in CaM expression are coupled to oncogene expression may yield insight into how CaM-regulated calcium response pathways are involved in mechanisms of oncogenic transformation.

Immunoreactive levels of myosin light-chain kinase in normal and virus-transformed chicken embryo fibroblasts.

Calmodulin, a calcium-modulated effector protein, is an important mediator of the intracellular actions of calcium through its interaction with calmodulin-binding proteins. We report here that the immunoreactive levels of a calmodulin-binding protein, myosin light-chain kinase, are decreased in transformed chicken embryo fibroblasts.

A knowledge-based experimental design system for nucleic acid engineering

Presented in this paper is a knowledge-based experimental design system that incorporates the domain expertise used in nucleic acid engineering, thus automating the processing of error-prone, laborious low-level work, and many decision-making steps, and guiding the biologist toward a workable plan. This allows the biologist to work at a higher abstraction level, concentrating on more fundamental, difficult and challenging problems directly related to protein structure - function relationships. Cassette-based site-directed mutagenesis and synthetic gene designs are used as examples to illustrate the utility of the knowledge-based system approach to experimental design.

Calcium Ions and the Dynamics of the Microtubular Cytoskeleton During the Initiation and the Progression of Mitosis in Endosperm Cells

Experiments have been carried out on endosperm cells of Amaryllidaceae (Haemanthus and Clivia) in order to investigate the role of Ca2+ as a second messenger in the regulation of the mitotic process.