Dicty News Electronic Edition Volume 17, number 14 December 15, 2001 Please submit abstracts of your papers as soon as they have been accepted for publication by sending them to dicty@northwestern.edu. Back issues of Dicty-News, the Dicty Reference database and other useful information is available at DictyBase--http://dictybase.org. =========================== New Report on DictyBase =========================== John Bonner has contributed a short report on the number of cells in Dictyostelium slugs. You can download this report from http://dictybase.org. ============= Abstracts ============= The Formation of Actin Rods Composed of Actin Tubules in Dictyostelium discoideum Spores Masazumi Sameshima,*1 Yoshiro Kishi,* Masako Osumi, Reiko Minamikawa-Tachino, Dana Mahadeo,2 and David A. Cotter2 *Electron Microscopy Center, and Computer Center, The Tokyo Metropolitan Institute of Medical Science, Tokyo Metropolitan Organization for Medical Research, 3-18-22 Honkomagome, Bunkyo-ku, Tokyo 113-8613, Japan; Department of Chemical and Biological Sciences, Faculty of Science, Japan Women's University, Bunkyo-ku, Tokyo 112-8681, Japan; and 2Department of Biological Sciences, University of Windsor, Windsor, Ontario N9B 3P4, Canada Accepted by J Struct Biol Abstract A new type of actin rods formed in both of the nucleus and the cytoplasm as well as tyrosine phosphorylation of actin are implicated in the maintenance of dormancy and viability of Dictyostelium discoideum spores. Recently we have found that S-adenosyl-L-homocysteine hydrolase that is a key enzyme for methylation is sequestered into actin rods. Here the ultrastructure of the rods and their relationship to the phosphorylation were examined. The rods first appeared in premature spores at the mid culmination stage as bundles composed of actin tubules hexagonally cross-linked. The 13 nm diameter bundles were composed of three actin filaments. Formation of the actin rods begins during late culmination stage and proceeds until two days after completion of fruiting bodies. The physical events occur in the following order; association of several modules of bundles, closely packing and decrease in diameter of actin tubules, elongation of rods across the nucleus or the cytoplasm. Actin phosphorylation levels increased at the late culmination stage and reached a maximum level 12 hours later. Immediately following activation of spore germination, actin was rapidly dephosphorylated, followed shortly thereafter with the disappearance of rods. Shortened actin tubules once again became arranged in a hexagonal pattern. This hexagonal arrangements of actin tubules are possibly involved in rod formation and disappearance, and do not depend upon actin phosphorylation. In contrast, rod-maturation processes may correlate with actin phosphorylation. ----------------------------------------------------------------------------- REGULATION OF ADENYLYL CYCLASES BY A REGION OUTSIDE THE MINIMALLY FUNCTIONAL CYTOPLASMIC DOMAINS Carole A. Parent, Jane Borleis, and Peter N. Devreotes Department of Biological Chemistry, The Johns Hopkins School of Medicine, Baltimore, MD 21205 Journal of Biological Chemistry, in press. The highly conserved topological structure of G protein activated adenylyl cyclases seems unnecessary since the soluble cytoplasmic domains retain regulatory and catalytic properties. Yet, we previously isolated a constitutively active mutant of the D. discoideum adenylyl cyclase harboring a single point mutation in the region linking the cytoplasmic and membrane domains (L394). We show here that multiple amino acid substitutions at L394 also display constitutive activity. The constitutive activity of these mutants is not dependent on G proteins or cytosolic regulators, although some of the mutants can be activated to higher levels than wild type. Combining a constitutive mutation such as L394T with K482N, a point mutation that renders the enzyme insensitive to regulators, restores an enzyme with wild type properties of low basal activity and the capacity to be activated by G proteins. Thus regions located outside the cytoplasmic loops of adenylyl cyclases are not only important in the acquisition of an activated conformation, they also have impact on other regions within the catalytic core of the enzyme. ----------------------------------------------------------------------------- Phospholipase Cd regulates germination of Dictyostelium spores Peter Van Dijken and Peter J.M. Van Haastert Department of Biochemistry, University of Groningen, Nijenborgh 4, 9747 AG Groningen, The Netherlands BMC Cell Biology, in press. Abstract Many eukaryotes, including plants and fungi make spores that resist severe environmental stress. The micro-organism Dictyostelium contains a single phospholipase C gene (PLC); deletion of the gene has no effect on growth, cell movement and differentiation. In this report we show that PLC is essential to sense the environment of food-activated spores. Plc-null spores germinate at alkaline pH, reduced temperature or increased osmolarity, conditions at which the emerging amoebae can not grow. In contrast, food-activated wild-type spores return to dormancy till conditions in the environment allow growth. The analysis of inositol 1,4,5-trisphosphate (IP3) levels and the effect of added IP3 uncover an unexpected mechanism how PLC regulates spore germination: i) deletion of PLC induces the enhanced activity of an IP5 phosphatase leading to high IP3 levels in plc-null cells; ii) in wild-type spores unfavourable conditions inhibit PLC leading to a reduction of IP3 levels; addition of exogenous IP3 to wild-type spores induces germination at unfavourable conditions; iii) in plc-null spores IP3 levels remain high, also at unfavourable environmental conditions. The results imply that environmental conditions regulate PLC activity and that IP3 induces spore germination; the uncontrolled germination of plc-null spores is not due to a lack of PLC activity but to the constitutive activation of an alternative IP3- forming pathway. ----------------------------------------------------------------------------- [End Dicty News, volume 17, number 14]