This is a proposal to identify and characterize membrane components of the rough endoplasmic reticulum and cytoplasmic factors which are involved in the recognition and binding of ribosomes and signal sequences in nascent polypeptide chains and may function as a vectorial discharge apparatus. Undenatured ribophorins and cytoplasmic protein of 93,000 daltons, will be purified by an affinity chromatography procedure based on the specific interaction of these proteins with the putative signal segments which are present in ovalbumin and in some mature membrane proteins, such as cytochrome P450 and epoxide hydratase. The affinity purified native proteins will be incorporated into the membranes of reconstituted vesicles which will be used in vitro assays to identify their functions in specific steps of the vectorial discharge. The effect of specific antiboides (monoclonal) against purified microsomal proteins on discrete steps of the cotranslational discharge of nascent polypeptides will be assessed in in vitro assays. Information on the structure and spatial dispositions of the ribophorins will be obtained using protein chemistry procedures, immunocytochemistry and sequencing of DNA's complementary to ribophorin mRNA's. Radioimmunoassays will be developed to establish correlations between the ribophorin content of microsomal membranes and their translocation activity during the development of the endoplasmic reticulum. The biosynthesis of ribophorins and the mechanisms for their insertion and retention in endoplasmic reticulum membranes will be studied using in vitro and in vivo systems. A possible role of the Golgi apparatus in the post-translational processing of these glycoproteins will be assessed.

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National Institute of General Medical Sciences (NIGMS)
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New York University
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Rajasekaran, A K; Langhans-Rajasekaran, S A; Gould, R M et al. (1995) A simple biochemical approach to quantitate rough endoplasmic reticulum. Am J Physiol 268:C308-16
Rajasekaran, A K; Morimoto, T; Hanzel, D K et al. (1993) Structural reorganization of the rough endoplasmic reticulum without size expansion accounts for dexamethasone-induced secretory activity in AR42J cells. J Cell Sci 105 ( Pt 2):333-45
Tsao, Y S; Ivessa, N E; Adesnik, M et al. (1992) Carboxy terminally truncated forms of ribophorin I are degraded in pre-Golgi compartments by a calcium-dependent process. J Cell Biol 116:57-67
Kelleher, D J; Kreibich, G; Gilmore, R (1992) Oligosaccharyltransferase activity is associated with a protein complex composed of ribophorins I and II and a 48 kd protein. Cell 69:55-65
De Lemos-Chiarandini, C; Ivessa, N E; Black, V H et al. (1992) A Golgi-related structure remains after the brefeldin A-induced formation of an ER-Golgi hybrid compartment. Eur J Cell Biol 58:187-201
Ivessa, N E; De Lemos-Chiarandini, C; Tsao, Y S et al. (1992) O-glycosylation of intact and truncated ribophorins in brefeldin A-treated cells: newly synthesized intact ribophorins are only transiently accessible to the relocated glycosyltransferases. J Cell Biol 117:949-58
Pirozzi, G; Zhou, Z M; D'Eustachio, P et al. (1991) Rat ribophorin II: molecular cloning and chromosomal localization of a highly conserved transmembrane glycoprotein of the rough endoplasmic reticulum. Biochem Biophys Res Commun 176:1482-6
Behal, A; Prakash, K; D'Eustachio, P et al. (1990) Structure and chromosomal location of the rat ribophorin I gene. J Biol Chem 265:8252-8
Yu, Y H; Sabatini, D D; Kreibich, G (1990) Antiribophorin antibodies inhibit the targeting to the ER membrane of ribosomes containing nascent secretory polypeptides. J Cell Biol 111:1335-42
Yu, Y H; Zhang, Y Y; Sabatini, D D et al. (1989) Reconstitution of translocation-competent membrane vesicles from detergent-solubilized dog pancreas rough microsomes. Proc Natl Acad Sci U S A 86:9931-5

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