CSM News Electronic Edition Volume 5, number 10 October 7, 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" =========== Abstracts =========== IDENTIFICATION OF TWO NOVEL Dictyostelium discoideum CYSTEINE PROTEINASES THAT CARRY GlcNAc-1-P MODIFICATION+ GLAUCIA M. SOUZA, JOHN HIRAI AND HUDSON H. FREEZE*. La Jolla Cancer Research Foundation, La Jolla, CA, 92037. J. Biol. Chem., in press. SUMMARY Dictyostelium discoideum makes multiple developmentally regulated lysosomal cysteine proteinases. One of these, a lysosomal enzyme called Proteinase I, contains a cluster of GlcNAc-a-1-P-Ser residues. We call this phosphoglycosylation. To study its function, a cDNA library from vegetative cells was screened and two novel cysteine proteinase clones were characterized (cprD and cprE). Each of them has highly conserved regions expected for cysteine proteinases, but unlike any other, each has a serine-rich domain containing three distinct motifs, poly-S, SGSQ and SGSG. cprD and cprE cDNAs were over-expressed in Dictyostelium and the active enzymes identified. cprD codes for a protein of approximately 36 kDa (CP4) which is recognized by monoclonal antibodies against GlcNAc-1-P and fucose. cprE corresponds to a 29 kDa protein which is recognized by antibodies against GlcNAc-1-P. mRNA for both enzymes is present in the vegetative phase and increases during growth on bacteria, but decreases throughout development. When the formation of the fruting body is complete the mRNA for both messages is detected again, but in very low levels. Having cloned cDNAs for proteins that carry GlcNAc-1-P should allow us to probe the function of the carbohydrate in this putative lysosomal enzymes. ----------------------------------------------------------------------- Expression of the human muscarinic receptor gene m2 in Dictyostelium discoideum Guido Voith, and Theodor Dingermann Institut for Parmaceutical Biology, Johann Wolfgang Goethe University, Biozentrum, Marie-Curie-Str. 9, D-60439 Frankfurt/Main, Germany BIO/TECHNOLOGY in press Summary The human muscarinic M2 receptor was functionally expressed in the cellular slime mold Dictyostelium discoideum, under the control of the homologous discoidin I promoter. The use of a contact site A leader peptide ensured insertion of the newly synthesized receptor protein into the plasma membrane.Due to the characteristics of the discoidin Ig promoter the M2 receptor is expressed late during growth and early during development. The heterologously expressed M2 receptors show binding characteristics similar to authentic receptors. Membranes as well as whole cells can be used in ligand binding assays. ---------------------------------------------------------------------- [End CSM News, volume 5, number 10]