Foot-and-mouth disease virus leader proteinase: purification of the Lb form and determination of its cleavage site on eIF-4 gamma

Abstract

Many picornaviruses cause a dramatic decrease in the translation of cellular mRNAs in the infected cell, without affecting the translation of their own RNA. Specific proteolysis of protein synthesis initiation factor eIF-4-y occurs during infection with rhinoviruses, enteroviruses, and aphthoviruses, apparently leading to an inability of the ribosomes to bind capped mRNAs. Cleavage of eIF-4-y in human rhinoviruses and enteroviruses is carried out by the viral 2A proteinase; in aphthoviruses (i.e., foot-and-mouth disease viruses), the leader proteinase is responsible for this reaction. We describe here the purification to homogeneity of the Lb form of the leader proteinase expressed in Escherichia coli. The primary cleavage products of eIF-4-y obtained in vitro with purified leader or 2A proteinase are electrophoretically indistinguishable from those found during infection in vivo. However, additional proteolysis products of eIF-4-y are observed with the leader proteinase and the human rhinovirus type 2 2A proteinase in vitro. The cleavage site of the leader proteinase in eIF-4y from rabbit reticulocyte was determined by sequencing the purified C-terminal cleavage product by automated Edman degradation. The cleavage site is between Gly-479 and Arg-480 and thus differs from that of rhinovirus and enterovirus 2A proteinases, which cleave between Arg-486 and Gly-487.

Foot-and-mouth disease virus induces lysosomal degradation of host protein kinase PKR by 3C proteinase to facilitate virus replication

“…levels after inhibition of cellular translational function, indicating that 3C pro -induced reduction of PKR protein levels is not totally related to the well-characterized blocking effect of 3C pro on cellular translation. L pro also induces the cleavage of host eIF4G to inhibit host protein synthesis (Kirchweger et al, 1994); however, we observed that over-expression of L pro had no detectable effect on PKR expression (Fig. 4A); it was similar to the observation in the cells treated with a translational inhibitor CHX (Fig.…”

Structure and Function of Viral Deubiquitinating Enzymes

“…Prior studies identified L pro as a potential virulence factor, with L pro cleaving the host eukaryotic translation initiation factor 4 gamma (eIF4G) that is involved in the translation of cellular and capped mRNA transcripts, thereby prioritizing the translation of viral transcripts [229]. Experiments using an FMDV mutant lacking the L pro domain also implicated the protease in the circumvention of cellular innate immune responses by the inhibition of cellular IFN-α/ β production and associated ISGs, specifically protein kinase R, a cytoplasmic sensor of viral RNA [230][231][232][233][234].…”

Translational control by viral proteinases

“…The 2A gene of FMDV is not a functional proteinase, rather cleavage of eIF4G is accomplished by the leader proteinase (L-pro), an alternate papain-like protease encoded at the N-terminus of the viral polyprotein. This proteinase cleaves eIF4GI at a distinct site, seven amino acids upstream of the 2A pro cleavage site, thereby accomplishing the same functional scis-sion of eIF4E-and eIF3-binding domains (Kirchweger et al, 1994). This cleavage reaction has been very well characterized and occurs extremely rapidly, presumably while ribosomes are translating the FMDV polyprotein (Glaser and Skern, 2000).…”