Evidence for a two-step mechanism involved in assembly of functional signal recognition particle receptor.
The signal recognition particle (SRP) and SRP receptor act sequentially to target nascent secretory proteins to the membrane of the ER. The SRP receptor consists of two subunits, SR alpha and SR beta, both tightly associated with the ER membrane. To examine the biogenesis of the SRP receptor we have developed a cell-free assay system that reconstitutes SR alpha membrane assembly and permits both anchoring and functional properties to be assayed independently. Our experiments reveal a mechanism involving at least two distinct steps, targeting to the ER and anchoring of the targeted molecule on the cytoplasmic face of the membrane. Both steps can be reconstituted in vitro to restore translocation activity to ER microsomes inactivated by alkylation with N-ethyl-maleimide. The characteristics elucidated for this pathway distinguish it from SRP-dependent targeting of secretory proteins, SRP-independent ER translocation of proteins such as prepromellitin, and direct insertion mechanisms of the type exemplified by cytochrome b5.
Andrews
D W
DW
Department of Physiology, University of California, San Francisco 94143.
Lauffer
L
L
Walter
P
P
Lingappa
V R
VR
D004721:Endoplasmic Reticulum / Q000378:metabolism*; D005033:Ethylmaleimide / Q000494:pharmacology; D008081:Liposomes / Q000378:metabolism; D008861:Microsomes / Q000378:metabolism*; D010196:Pancreatic Elastase / Q000378:metabolism; D011388:Prolactin / Q000378:metabolism; D011498:Protein Precursors / Q000378:metabolism; D011956:Receptors, Cell Surface / Q000096:biosynthesis / Q000378:metabolism*; D018160:Receptors, Cytoplasmic and Nuclear; D018000:Receptors, Peptide; D012261:Ribonucleoproteins / Q000378:metabolism; D018271:Signal Recognition Particle; D014357:Trypsin / Q000378:metabolism
D008081:Liposomes; D011498:Protein Precursors; D011956:Receptors, Cell Surface; D018160:Receptors, Cytoplasmic and Nuclear; D018000:Receptors, Peptide; D012261:Ribonucleoproteins; D018271:Signal Recognition Particle; C036318:signal peptide receptor; C017671:preprolactin; D011388:Prolactin; D010196:Pancreatic Elastase; D014357:Trypsin; D005033:Ethylmaleimide