Specific types of prosomes distribute differentially between intermediate and actin filaments in epithelial, fibroblastic and muscle cells Cristina Arcangeletti 2)a,b , Flora De Conto 2)a,b , Rosmarie Sütterlin c , Federica Pinardi a,b , Silvia Missorini a,b , Gerard Ge  raud a , Ueli Aebi c , Carlo Chezzi b , Klaus Scherrer 1)a a Institut J. Monod, CNRS, Universite  Paris 7, Paris/France b Sezione Microbiologia, Dipartimento Patologia e Medicina di Laboratorio, Universita Á degli Studi di Parma, Parma/ Italy c M. E. Müller Institute, Biozentrum, University Basel, Basel/Switzerland Received May 11, 1999 Received in revised version January 3, 2000 Accepted January 18, 2000 Prosomes ± proteasomes ± mRNP ± intermediate filaments ± microfilament-stress fibers ± actin First observed as components of non-translated mRNP com- plexes, prosomes harbour RNase and several proteinase activities; they are also the central constituent of the ªMulti- catalytic Proteinase (MCP) complexesº or ª26S-protea- somesº. In two recent publications (Arcangeletti et al., 1997b; De Conto et al., 1997) we have shown, by applying a new fixation technique, that these particles distribute differ- entially between the cytoskeletal networks of intermediate filament (IF) and actin types; previously they had been observed exclusively on the intermediate filaments. Here we further investigate the distribution of prosomes of several types, distinct by their subunit composition, between the IF of vimentin type and the actin network, as well as in the 3D space of the cell. It is shown that subtypes of prosomes occupy specific networks of the cytoskeleton, and that this pattern is specific for a given cell type. Confocal microscopy shows that prosome cytodistribution is not homogeneous in the 3D space: in the perinuclear area they colocalize most strongly with the IF, and more peripherally with the microfilament/stress fiber system; connections may exist between the two networks. Furthermore, new data indicate that the prosome-actin inter- action may participate in the molecular structure of the stress fibers. Definitions and Abbreviations. We use here the term ªProsomesº introduced by our laboratory (Schmid et al., 1984) for the then unknown mRNA-associated 19 ± 20S particle. When its proteinase activity is addressed, the term ªMulticatalytic Proteinase or MCPº was recommended by the group of enzymologists concerned (Baumeister et al., 1988; Orlowski and Wilk, 1988), in preference to the term ªProteasomeº suggested by Arrigo et al. (Arrigo et al., 1988). The term ªProsomesº relates to the pre-translational functions of the 20S particles, mRNA- and cytoskeleton-related, and ªProteasomesº to its catabolic activity carried out, in vivo, exclusively within the 26S proteasome complexes (Baumeister et al., 1998; Larsen and Finley, 1997). ± ABP Actin-binding protein. ± CSK Cytoskeleton. ± EM Electron microscopy. ± IIF Indirect immunofluores- cence. ± IFs Intermediate filaments. ± MCP Multicatalytic proteinase. ± MFs Microfilaments. ± MTs Microtubules. ± p-mAb Anti-prosome monoclonal antibody. ± p25K, p27K, p29K, p30K, p31K Prosomal subunit proteins of Mr 25 000, 27 000, 29 000, 30 000, 31 000, respectively. Introduction The topology of cytodistribution of specific components of the cell is a classical aspect of cell biology. More recently, cytolocalization has also become a major aspect of molecular biology, and in particular of gene expression by protein biosynthesis. Indeed, data acquired over the last ten years show, not only that genes occupy specific sectors of interphase nuclei, but also that specific types of mRNA are transported to highly specific sectors of the cytoplasm (Bassell et al., 1994; Hesketh, 1991; Hesketh, 1996; Kislauskis et al., 1994; Singer, 1992). An unambiguous demonstration of this fact is the intercalation of specific mRNAs right into the sarcomeric pattern of skeletal muscle indicating, possibly, co-translational assembly of the actin-myosin complex (Fulton, 1993). Re- cently we found that specific types of prosomes form a similar pattern in skeletal muscle cells (Foucrier et al., 1999; Grand et al., 1994). Furthermore, we have shown that some types of prosomes align exclusively along the bile canaliculi, there where the bile-specific proteins are being produced (Briane et al., 1992). The cytoskeleton seems to be involved in prosome cytolocalization (see discussion in (Scherrer and Bey, 1994)) but also of mRNA: messenger RNA was found on the actin system (Singer, 1992) and was also observed, more recently, on EJCB 423 European Journal of Cell Biology 79, 423 ± 437 (2000, June) ´  Urban & Fischer Verlag ´ Jena http://www.urbanfischer.de/journals/ejcb 0171-9335/00/79/06-423 $12.00/0 1) Prof. Dr. K. Scherrer, Institut Jacques Monod, CNRS and Universite  Paris 7, 2, place Jussieu, F-75251 Paris Cedex 05/France, e-mail: scherrer@ijm.jussieu.fr, Fax:  33 1 44 27 76 47. 2) These authors contributed equally to this investigation.