Comparative processing of bovine leukemia virus envelope glycoprotein gp72 by subtilisin/kexin‐like mammalian convertases

Abstract

Intracellular proteolytic processing of bovine leukemia virus (BLV) envelope glycoprotein precursor (gp72) at the Cterminal end of the RVRR 268 J, site is an essential step for virus infectivity. Subtilisin/kexin-like convertases cleave proproteins at preferred RX(K/R)R J, sites, including those commonly found in viral envelope glycoprotein precursors. We first demonstrated that gp72 is processed into gp51/gp30 in both CV1 cells and the furin-deficient LoVo cells, leading us to compare the ability of mammalian convertases to cleave BLV gp72 in vitro. In contrast to the inability of the neuroendocrine PCI to cleave gp72, the convertases furin, PACE4, PC5-A and PC5-B, which process constitutively secreted precursors, can effectively cleave gp72 into gp51/gp30. N-terminal sequence analysis of the convertasegenerated gp30 demonstrated that cleavage occurs at the in vivoutilized RVRRiSPV site. Such furin-, PACE4-and PC5-mediated processing was completely inhibited by the oiiantitrypsin variant od-PDX. Mutagenesis of the gp72 cleavage site into RVRG-TPV resulted in complete abrogation of gp72 processing by endogenous CV-1 cells and by convertases in vitro. Since our in vitro data suggest a redundancy in the ability of the convertases to cleave gp72, RT-PCR analysis was used to define the convertases expressed in B-lymphocytes, representing one of the major targets of BLV infection. Our data revealed that only furin and the newly discovered PC7 mRNAs are expressed in Raji, B-Jab and LG2 cell lines.

Regulation of Membrane Type-1 Matrix Metalloproteinase Activation by Proprotein Convertases

“…As expected, neither of the well-characterized furin-deficient cell lines, LoVo or RPE.40, processed soluble ⌬MT1-MMP to its mature form. LoVo cells have been reported to express PACE4 as well as PC7, and this furin-deficient cell line can process other RXKRencrypted targets including human immunodeficiency virus-1, E-cadherin and gp-160 (Gu et al, 1995;Decroly et al, 1997;Zarkik et al, 1997;Posthaus et al, 1998). However, neither PACE4 nor PC7 appeared to play a major role in the processing of ⌬MT1-MMP in LoVo cells.…”

“…However, in contrast to LoVo cells, RPE.40 cells do express PC6 (as both the transmembrane-anchored and soluble forms, PC6B and P6CA, respectively; Duguay et al, 1997). Interestingly, PC6, like furin, can process RXKR-, RXXR-, and KXXR-containing molecules (Zarkik et al, 1997;Cui et al, 1998). Furthermore, because PC6 cannot be de-tected in LoVo cells (Vollenweider et al, 1996;Logeat et al, 1998), differences in the MT1-MMP processing pathways displayed between LoVo and RPE.40 cells are likely attributed to each cell line's distinct repertoire of processing enzymes.…”

Subtilase-like pro-protein convertases: from molecular specificity to therapeutic applications

“…It also behaves as a suicide substrate inhibitor, as binding to the active site results in an equal probability that proteolysis will ensue or that a kinetically trapped SDS-stable complex with the enzyme will be formed (Dufour et al 1998. Because of its high selectivity toward SPC1 , this novel inhibitor has already been used to assess the biological role of SPC1 in processing different precursors , Denault et al 1995b, Watanabe et al 1995, Decroly et al 1996, Kayo et al 1997, Zarkik et al 1997, Abrami et al 1998, Cui et al 1998, Logeat et al 1998. Other naturally occuring substances have been tested for their ability to inhibit SPCs.…”

Implication of the proprotein convertases furin, PC5 and PC7 in the cleavage of surface glycoproteins of Hong Kong, Ebola and respiratory syncytial viruses: a comparative analysis with fluorogenic peptides

“…The separation was monitored on-line by both UV absorbance at 210 nm and fluorescence (λ ex l 320 nm and λ em l 420 nm), measured in arbitrary units. The various peaks with retention times shown in parenthesis were analysed by amino acid composition and MALDI-TOF-MS. Abbreviations : NT 1 , fluorescent N-terminal fragment (1-13) generated by cleavage at the physiological site (heavy arrow) ; NT 2 , fragment (1-9) generated by cleavage at an alternate site (thin arrow) ; CT 1 and CT 2 , non-fluorescent C-terminal fragments (14)(15)(16)(17) and (10)(11)(12)(13)(14)(15)(16)(17) repectively.…”

“…Thus it has been demonstrated that conditions favouring viral replication cause a significant upregulation of candidate processing enzymes such as furin and PC7 [14][15][16][17][18]. These and other studies have indicated that furin and PC7 are the major candidate surface glycoprotein (including HIV gp160) convertases in lymphocytes [15][16][17][18]. This finding inspired various groups to examine the possible participation of PCs in other viral and opportunistic infections [19][20][21][22].…”