TheYeastAcidPhosphatase CanEntertheSecretory Pathway without ItsN-Terminal Signal Sequence

Therepressible Saccharomyces cerevisiae acidphosphatase (APase) codedbythePHOSgeneisacell wall glycoprotein thatfollows theyeast secretory pathway. We usedinvitro mutagenesis toconstruct adeletion (ASP)including theentire signal sequence andfouraminoacids ofthematuresequence ofAPase.An APase-deficient yeast strain wastransformed withahigh-copy-number plasmid carrying thePHOS/ASPgene. Whenexpressed invivo, thePHO5/ASP geneproduct accumulated predominantly asaninactive, unglycosylatedformlocated inside thecell. A large partofthisunglycosylated precursor underwent proteolytic degradation, butupto30%ofitwastranslocated, coreglycosylated, andmatured bytheaddition ofmannose residues, before reaching thecell wall. Itappears, therefore, that thesignal sequence isimportant forefficient translocation andcoreglycosylation ofyeast APasebutthat itisnotabsolutely necessary forentry oftheprotein into theyeast secretory pathway. mRNA obtained byinvitro transcription ofPHOSandPHO5/ASP genes were translated invitro inthepresence ofeither reticulocyte lysate anddogpancreatic microsomes oryeast lysate andyeast microsomes. ThePHO5geneproduct wastranslocated andcoreglycosylated intheheterologous system andlessefficiently inthehomologous system. We werenotabletodetect anytranslocation or glycosylation ofPHO51ASP geneproduct intheheterologous system, buta verysmallamountofcore suppression ofglycosylated material could beevidenced inthehomologous system. Mosteucaryotic andprocaryotic secretory proteins are synthesized invitro asprecursors withN-terminal exten