Identification of Lassa virus glycoprotein signal peptide as a trans‐acting maturation factor

Abstract

Lassa virus glycoprotein is translated as a precursor (pre-GP-C)into the lumen of the endoplasmic reticulum and is cotranslationally cleaved into the signal peptide and GP-C, before GP-C is proteolytically processed into its subunits GP1 and GP2. The signal peptide of pre-GP-C comprises 58 amino acids. The substitution of Lassa virus pre-GP-C signal peptide with another signal peptide still mediates translocation and the release of signal peptide but abolishes the proteolytic cleavage of GP-C into GP1 and GP2. Remarkably, cleavage of GP-C from these hybrid pre-GP-C substrates was restored on coexpression of the wildtype pre-GP-C signal peptide, indicating that the signal peptide functions as a trans-acting factor to promote Lassa virus GP-C processing. To our knowledge, this is the first report on a signal peptide that is essential for proteolytic processing of a secretory pathway protein.

Maturation cleavage within the ectodomain of Lassa virus glycoprotein relies on stabilization by the cytoplasmic tail

“…We performed sucrose gradient centrifugation as described above of cells expressing a LASV GP mutant which contains the signal peptide of an influenza hemagglutinin protein (LASV GP-C/HA-SP). This mutant lacks its signal peptide since it has been previously shown that the signal peptide of HA does not stay connected to the LASV GP complex after signal peptidase processing and does not support the cleavage of GP-C into GP-1 and GP-2 [3]. During sucrose gradient centrifugation the mutated proteins migrated in the same way as wild type LASV GP (Fig.…”

“…One characteristic of LASV is that the signal peptide acts as an essential factor during maturation cleavage of the LASV glycoprotein [3], by a still unknown mechanism. To exclude any impact of the SSP on the results we observed in this work we investigated the contribution of the SSP on the oligomerization and stability of the glycoprotein complex.…”

“…The latter one includes the prototypic family members lymphocytic choriomeningitis virus (LCMV) and Lassa virus (LASV) [1]. The glycoprotein of LASV is synthesized as the precursor molecule preGP-C which is co-translationally processed by the cellular signal peptidase resulting in the 6 kDa stable signal peptide (SSP) and the immature 76 kDa GP-C. Uncommonly, after cleavage the SSP stays connected to the glycoprotein complex [2][3][4]. During the transport along the secretory pathway to the plasma membrane, GP-C is then further proteolytically activated by maturation cleavage into the distal glycoprotein subunit (GP-1) (44 kDa) and the transmembrane-spanning subunit (GP-2) (36 kDa) by the cellular endoprotease subtilase subtilisin kexin isozyme-1/site 1 protease (SKI-1/S1P) [5,6].…”

A Novel Zinc-Binding Domain Is Essential for Formation of the Functional Junín Virus Envelope Glycoprotein Complex

“…GP-C from the pathogenic JUNV strain MC2 (24) was expressed in Vero cells by cotransfection of pcDNA3.1 (Invitrogen) plasmids encoding CD4sp-GPC (in which SSP is replaced by the conventional signal peptide of CD4) and SSP-term (in which a stop codon is introduced following the C-terminal SSP amino acid T58) (65). These components associate in trans and reconstitute the native GP-C complex (15). Transient expression utilized a recombinant vaccinia virus expressing T7 RNA polymerase (vTF7-3 [20]) and the T7 promoter of pcDNA3.1.…”

“…The stable, 58-residue signal peptide (SSP) was initially identified as a component of the Old World arenavirus GP-C complex (16,19,35) and found to be essential for proteolytic maturation of the glycoprotein precursor in the Golgi body (1,15,35). Without SSP, the G1-G2 precursor is specifically retained in the endoplasmic reticulum (ER), in part through dibasic ER retrieval signals in the cytoplasmic domain (CTD) of G2 (1).…”