CSM News Electronic Edition Volume 5, number 2 July 15, 1995 Please submit abstracts of your papers as soon as they have been accepted for publication by sending them to CSM-News@worms.cmsbio.nwu.edu. Back issues of CSM-News, the CSM Reference database and other useful information is available by anonymous ftp from worms.cmsbio.nwu.edu [165.124.233.50], via Gopher at the same address, or by World Wide Web at the URL "http://worms.cmsbio.nwu.edu/dicty.html" ==================== Announcing Dicty96 ==================== Dear Dicty people, The next International Dictyostelium Conference'96 is scheduled to be held in Sendai, Japan, from October 13th(Sun) to October 18th(Fri), 1996. Sendai City is located northeast about 400km from Tokyo. The city is not so big, not so small, and one of the most beautiful cities in Japan, which is famous for "Sendai virus" and is called "a tree-clad town". The place of meeting (Sendai City Hole) is just in the down-town of Sendai, and therefore you will be able to see several streets with a lot of nicely arranged Japanese Zelkovas, shops, and restraunts including "Karaoke bars" around there. During the period of meeting (maybe evening of October 16th, 1996), we are planning to go to a hot spring resort near Sendai and stay overnight in a nice hotel of purely Japanese style to have an "extra" session. Fortunately, the meeting will be partly supported by a fund from Yamada Science Foundation, Japan. As you know well, however, Japanese currency (yen) is now extremly high, though we Japanese are not necessarily rich in Japan. By the way, to welcome many people to the meeting, we are seriously hoping that Japanese yen will become lower and lower. I will occasionally inform you the details of the meeting hereafter. Again, I would like to express many thanks to Drs. Michael Veron and Michel Satre for organizing nicely the Dicty'95 meeting in France and for providing a lot of delicious wine. Hoping to see you next year in Japan. Yasuo Maeda, Organizer of the International Dictyostelium Conference'96. Biological Institute, Graduate School of Science, Tohoku University, Aoba, Sendai 980-77, Japan. TEL & FAX: +81-22-217-6709 E-mail: j23023@cctu.cc.tohoku.ac.jp -------------------------------------------------------------------------- =========== Abstracts =========== IDENTIFICATION OF MAJOR PROTEINS ASSOCIATED WITH DICTYOSTELIUM DISCOIDEUM ENDOCYTIC VESICLES Celine Adessi, Agnes Chapel, Mathilde Vincon, Thierry Rabilloud, Gerard Klein, Michel Satre and Jerome Garin Departement de Biologie Moleculaire et Structurale, CEN-G, 17 rue des Martyrs, 38054 Grenoble Cedex 09, France. J. Cell Sci., in press SUMMARY Magnetic isolation of endocytic vesicles from Dictyostelium discoideum was accomplished after feeding the amoebae with iron oxide particles. Proteins associated with the endocytic vesicles were resolved by SDS-PAGE and digested "in-gel" with endoproteinase Lys-C or Asp-N to generate peptides for amino acid sequencing. This strategy allowed the identification of the major protein constituents of the vesicles, namely the A, B, D, E and 110 kDa subunits of a vacuolar type H+-ATPase, actin, a Rab 7-like GTPase, a p34 protein corresponding to a new cysteine proteinase and the 25 kDa product of a recently sequenced D. discoideum open reading frame (Winckler, T., 1995; GeneBank U20997). ---------------------------------------------------------------------- AT-Rich Upstream Sequence Elements Regulate Spore Germination-Specific Expression of the Dictyostelium discoideum celA Gene Ramachandran Ramalingam, John E. Blume,1 Kalyan Ganguly2 and Herbert L. Ennis Roche Institute of Molecular Biology, 1 Sandoz Research Institute, 2 Department of Anatomy and Cell Biology, State University of New York Health Science Center at Brooklyn Nucleic Acids Research, in press Summary Two members of a family of spore germinationsp ecific cDNAs, celA and celB are expressed coordinately, exclusively during spore germination. In the present study the regulatory sequence elements responsible for celA's germination specific expression have been identified. The very AT rich 81 bp sequence between -664 and -584 upstream of the translation initiation site was required for the proper temporal transcription of the celA gene. This sequence was comprised of two cis elements, each of which was active by itself in allowing celA expression. Electrophoretic mobility shift assays showed that factor(s) in an extract prepared from germinating spores bound to the celA regulatory region. One of the three complexes formed was specific for the germinating spore extract. The results are consistent with the notion that the factor(s) that binds to this regulatory region are involved in expression of celA. ----------------------------------------------------------------------- A Phosphatidylinositol Kinase Gene Family in Dictyostelium discoideum: Biological Roles of Putative Mammalian p110 and Yeast Vps34p PI 3-Kinase Homologs During Growth and Development Kemin Zhou1, Kaoru Takegawa2, Scott D. Emr2, and Richard A. Firtel1 1Department of Biology, Center for Molecular Genetics and 2Division of Cellular and Molecular Medicine, School of Medicine, Center for Molecular Genetics, Howard Hughes Medical Institute University of California, San Diego 9500 Gilman Drive, La Jolla, CA 92093-0634 Mol. Cell. Biol., in press ABSTRACT Three groups of phosphotidylinositol kinases (PI kinase) convert phosphatidylinositol (PI) into PI(3)phosphate [PI(3)P], PI(4)phosphate [PI(4)P], PI(4,5)bisphosphate [PI(4,5)P2], and PI(3,4,5)trisphosphate [PI(3,4,5)P3]. These phosphoinositides have been shown to function in vesicle-mediated protein sorting and they serve as second messenger signaling molecules for regulating cell growth. To further elucidate the mechanism of regulation and function of phosphoinositides, we cloned genes encoding five putative PI kinases from Dictyostelium discoideum. Data base analysis indicates that DdPIK1, 2, and 3 are most closely related to the mammalian p110 PI 3-kinase, DdPIK5 is closest to the yeast Vps34 PI 3-kinase, and DdPIK4 is most homologous to PI 4-kinases. Together with other known PI kinases, a superfamily of PI kinase genes has been defined with all the encoded proteins sharing a common highly conserved catalytic core domain. DdPIK1, 2, and 3 may have redundant functions because disruption of any single gene had no effect on D. discoideum growth or development. However, strains in which both of the two most highly related genes, DdPIK1 and DdPIK2, were disrupted showed both growth and developmental defects, while double knockouts of DdPIK1 and DdPIK3 and DdPIK2 and DdPIK3 appear to be lethal. The Ddpik1 Ddpik2 null cells were smaller than wild-type cells and grew slowly both in association with bacteria and in axenic medium when attached to Petri plates but were unable to grow in suspension in axenic medium. When DDdpik1 DDdpik2 null cells were plated for multicellular development, they formed aggregates having multiple tips and produced abnormal fruiting bodies. Antisense expression of DdPIK5 (a putative homolog of the S. cerevisiae VPS34) led to a defect in the growth of D. discoideum cells on bacterial lawns and abnormal development. DdPIK5 complemented the temperature-sensitive growth defect of a Schizosaccharomyces pombe DSpvps34 mutant strain, suggesting DdPIK5 encodes a functional homolog of the yeast Vps34 protein. These observations indicate that, in D. discoideum, different PI kinases regulate distinct cellular processes, including cell growth, development, and protein trafficking. ------------------------------------------------------------------------ [End CSM-News, volume 5, number 2]