Facultative polypeptide translocation allows a single mRNA to encode the secreted and cytosolic forms of plasminogen activators inhibitor 2.

Abstract

Two forms of plasminogen activators inhibitor 2 (PAI‐2) are synthesized by human and murine monocytes/macrophages: one accumulates in the cytosol, while the other is translocated into the endoplasmic reticulum, glycosylated and secreted. We show here that a single mRNA encodes both forms of PAI‐2. Firstly, a single PIA‐2 mRNA was detected by Northern blot hybridization and by RNase protection. Secondly, transfection of a PAI‐2 cDNA led to the synthesis of both forms of PAI‐2. Finally, in vitro translation of an mRNA transcript of the PAI‐2 cDNA in the presence of microsomal membranes generated two topologically distinct forms of PAI‐2. The cytosolic and secreted forms of PAI‐2 do not result from the use of two translation start sites, since their synthesis initiates at the same AUG, in a sequence context that is conserved between the human and murine genes. Thus, the accumulation of one polypeptide into two topologically distinct cellular compartments can be achieved by facultative translocation.

Protease-nexin I as an androgen-dependent secretory product of the murine seminal vesicle.

“…The blots were hybridized at 58°C, and the three stringency washes were done at 760C. RNase protection assays and in situ hybridizations were performed as described by Belin et al (1989) and Sappino et al (1991). A 1 kbp murine PN-I probe, obtained by PCR amplification of seminal vesicle cDNA using two oligonucleotides corresponding to regions conserved between rat and human PN-I (Sommer et al, 1987), was cloned into pGEM-3Z and transcribed with SP6 RNA polymerase; the probe corresponds to the sequence coding for 11e42 to Ser366 (Sommer et al, 1987).…”

Coupled Translocation Events Generate Topological Heterogeneity at the Endoplasmic Reticulum Membrane

“…Specialized topogenic determinants in naturally occurring proteins may also direct alternate topologies. In the case of murine plasminogen activator inhibitor 2, cytosolic and secretory iso-forms are generated by a bipartite signal sequence that targets the nascent chain to the ER membrane but initiates translocation for only a fraction of targeted chains (Belin et al, 1989(Belin et al, , 1996. Similarly, secretory and TM conformations of the prion protein (Hay et al, 1987a,b) PrP are directed by a specialized pause transfer sequence that transiently terminates translocation and allows a downstream hydrophobic segment to span the membrane in a subset of nascent chains Nakahara et al, 1994).…”

“…Present address: Division of Molecular Medicine, Oregon Health Sciences University, Portland, OR 97201-3098. 1 Abbreviations used: ER, endoplasmic reticulum; MDR1-Pgp, human P-glycoprotein; Pgp, P-glycoprotein; PK, proteinase K; ST, stop transfer; TM, transmembrane; XO, Xenopus oocyte.© 1998 by The American Society for Cell Biology 2681 forms are generated by a bipartite signal sequence that targets the nascent chain to the ER membrane but initiates translocation for only a fraction of targeted chains (Belin et al, 1989(Belin et al, , 1996. Similarly, secretory and TM conformations of the prion protein (Hay et al, 1987a,b) PrP are directed by a specialized pause transfer sequence that transiently terminates translocation and allows a downstream hydrophobic segment to span the membrane in a subset of nascent chains Nakahara et al, 1994).…”

39 Structure function relationships of plasminogen activator inhibitor type-2 (PAI-2)

“…Both the 47kD (primarily intracellular) and 60kD (glycosylated/secreted) forms of PAI-2 are encoded by a single mRNA, the bi-topological distribution of PAI-2 resulting from facultative translocation into the endoplasmic reticulum (Belin et al 1989). This process appears to be under the control of an uncleaved internal secretion signal, similar to that found in ovalbumin, between residues 4-16 and 25-46.…”

“…Regions of low sequence homology among different serpins are found in a range of N-terminal extensions to the core domain. These extensions serve as cleavable signal peptides in many secreted members of the family or as an internal secretion signal sequence in PAI-2 and ovalbumin (which both lack cleavable signal peptides) (Belin et al 1989;Belin 1993). Other extensions include N-terminal region in antithrombin III and PAI-1 that facilitate enhanced inhibitory function via hepar binding (Gettins et al 1996), and a proline and serine rich N-terminal extension in inhibitor, allowing heavy glycosylation (Schoenberger 1992).…”