scholarly journals Sequential modification of translation initiation factor eIF4GI by two different foot-and-mouth disease virus proteases within infected baby hamster kidney cells: identification of the 3Cpro cleavage site

2004 ◽  
Vol 85 (10) ◽  
pp. 2953-2962 ◽  
Author(s):  
Rebecca Strong ◽  
Graham J. Belsham
Keyword(s):  
Amino Acid ◽  
Disease Virus ◽  
Translation Initiation ◽  
Cleavage Site ◽  
Foot And Mouth Disease ◽  
Translation Initiation Factor ◽  
Initiation Factor ◽  
Baby Hamster Kidney ◽  
3C Protease ◽  
Mouth Disease Virus

Infection of cells by foot-and-mouth disease virus (FMDV) causes the rapid inhibition of cellular cap-dependent protein synthesis that results from cleavage of the translation initiation factor eIF4G, a component of the cap-binding complex eIF4F. Two FMDV proteins, the leader (L) and 3C proteases, have been shown individually to induce cleavage of eIF4GI at distinct sites within baby hamster kidney (BHK) cells. Here, sequential cleavage of eIF4GI by the L and 3C proteases was demonstrated in FMDV-infected BHK cells. The FMDV 3C cleavage site within hamster eIF4GI was localized to a small region (about 40 aa) of the protein, between the sites cleaved by the poliovirus 2A protease and the human immunodeficiency virus type 2 protease. Human eIF4GI was found to be resistant to the action of the FMDV 3C protease. On the basis of amino acid sequence alignments, it was predicted and then verified that substitution of a single amino acid residue within this region of human eIF4GI conferred sensitivity to cleavage by the FMDV 3C protease within cells. Full-length eIF4GI and both forms of the C-terminal cleavage product must be capable of supporting the activity of the FMDV internal ribosome entry site in directing translation initiation.

scholarly journals Foot-and-Mouth Disease Virus 3C Protease Induces Cleavage of Translation Initiation Factors eIF4A and eIF4G within Infected Cells

2000 ◽  
Vol 74 (1) ◽  
pp. 272-280 ◽  
Author(s):  
Graham J. Belsham ◽  
Gerald M. McInerney ◽  
Natalie Ross-Smith
Keyword(s):  
Disease Virus ◽  
Translation Initiation ◽  
Foot And Mouth Disease ◽  
Mouth Disease ◽  
Translation Initiation Factor ◽  
Initiation Factor ◽  
Cell Protein ◽  
3C Protease ◽  
Mouth Disease Virus ◽  
Foot And Mouth

ABSTRACT Infection of cells by foot-and-mouth disease virus (FMDV) results in the rapid inhibition of host cell protein synthesis. This process is accompanied by the early cleavage of the translation initiation factor eIF4G, a component of the cap-binding complex eIF4F. This cleavage is mediated by the leader (L) protease. Subsequently, as the virus proteins accumulate, secondary cleavages of eIF4G occur. Furthermore, eIF4A (46 kDa), a second component of eIF4F, is also cleaved in these later stages of the infection cycle. The 33-kDa cleavage product of eIF4A has lost a fragment from its N terminus. Transient-expression assays demonstrated that eIF4A was not cleaved in the presence of FMDV L or with the poliovirus 2A protease (which also mediates eIF4G cleavage) but was cleaved when the FMDV 3C protease was expressed. The FMDV 3C protease was also shown in such assays to induce cleavage of eIF4G, resulting in the production of cleavage products different from those generated by the L protease. Consistent with these results, within cells infected with a mutant FMDV lacking the L protease or within cells containing an FMDV replicon lacking L-P1 coding sequences it was again shown that eIF4A and eIF4G were cleaved.

scholarly journals Functional interaction of translation initiation factor eIF4G with the foot-and-mouth disease virus internal ribosome entry site

2001 ◽  
Vol 82 (4) ◽  
pp. 757-763 ◽  
Author(s):  
Lanja Saleh ◽  
René C. Rust ◽  
Ralf Füllkrug ◽  
Ewald Beck ◽  
Gergis Bassili ◽  
...  
Keyword(s):  
Translation Initiation ◽  
Foot And Mouth Disease ◽  
Translation Initiation Factor ◽  
Initiation Factor ◽  
Entry Site ◽  
Stem Loop ◽  
Internal Ribosome Entry ◽  
Initiation Factors ◽  
Mouth Disease Virus ◽  
Ires Element

In the life-cycle of picornaviruses, the synthesis of the viral polyprotein is initiated cap-independently at the internal ribosome entry site (IRES) far downstream from the 5′ end of the viral plus-strand RNA. The cis-acting IRES RNA elements serve as binding sites for translation initiation factors that guide the ribosomes to an internal site of the viral RNA. In this study, we show that the eukaryotic translation initiation factor eIF4G interacts directly with the IRES of foot-and-mouth disease virus (FMDV). eIF4G binds mainly to the large Y-shaped stem–loop 4 RNA structure in the 3′ region of the FMDV IRES element, whereas stem–loop 5 contributes only slightly to eIF4G binding. Two subdomains of stem–loop 4 are absolutely essential for eIF4G binding, whereas another subdomain contributes to a lesser extent to binding of eIF4G. At the functional level, the translational activity of stem–loop 4 subdomain mutants correlates with the efficiency of binding of eIF4G in the UV cross-link assay. This indicates that the interaction of eIF4G with the IRES is crucial for the initiation of FMDV translation. A model for the interaction of initiation factors with the IRES element is discussed.

scholarly journals Interaction of Eukaryotic Initiation Factor eIF4B with the Internal Ribosome Entry Site of Foot-and-Mouth Disease Virus Is Independent of the Polypyrimidine Tract-Binding Protein

1999 ◽  
Vol 73 (7) ◽  
pp. 6111-6113 ◽  
Author(s):  
René C. Rust ◽  
Kerstin Ochs ◽  
Karsten Meyer ◽  
Ewald Beck ◽  
Michael Niepmann
Keyword(s):  
Disease Virus ◽  
Translation Initiation ◽  
Internal Ribosome Entry Site ◽  
Foot And Mouth Disease ◽  
Initiation Factor ◽  
Polypyrimidine Tract ◽  
Entry Site ◽  
Internal Ribosome Entry ◽  
Polypyrimidine Tract Binding Protein ◽  
Mouth Disease Virus

ABSTRACT Eukaryotic translation initiation factor 4B (eIF4B) binds directly to the internal ribosome entry site (IRES) of foot-and-mouth disease virus (FMDV). Mutations in all three subdomains of the IRES stem-loop 4 reduce binding of eIF4B and translation efficiency in parallel, indicating that eIF4B is functionally involved in FMDV translation initiation. In reticulocyte lysate devoid of polypyrimidine tract-binding protein (PTB), eIF4B still bound well to the wild-type IRES, even after removal of the major PTB-binding site. In conclusion, the interaction of eIF4B with the FMDV IRES is essential for IRES function but independent of PTB.

Caspases are not involved in the cleavage of translation initiation factor eIF4GI during picornavirus infection

Microbiology ◽  
2000 ◽  
Vol 81 (7) ◽  
pp. 1703-1707 ◽  
Author(s):  
Lisa O. Roberts ◽  
Angela J. Boxall ◽  
Louisa J. Lewis ◽  
Graham J. Belsham ◽  
George E. N. Kass
Keyword(s):  
Translation Initiation ◽  
Caspase 3 ◽  
Foot And Mouth Disease ◽  
Cellular Protein ◽  
Translation Initiation Factor ◽  
Initiation Factor ◽  
Fmdv Infection ◽  
Expression Studies ◽  
Mouth Disease Virus ◽  
Picornavirus Infection

Infection of cells by many picornaviruses results in the rapid inhibition of cellular protein synthesis due to cleavage of the translation initiation factor eIF4G. The poliovirus (PV) 2A and foot-and-mouth disease virus (FMDV) L proteases are each sufficient to mediate this cleavage, but the cleavage mechanism may be indirect, involving an unidentified cellular protease(s). eIF4G is also targetted for cleavage by caspase-3 during apoptosis. Here, it is shown that caspase inhibitors do not inhibit the cleavage of eIF4GI during PV or FMDV infection. Similarly, in transient-expression studies, the cleavage of eIF4GI induced by PV 2A or FMDV L was unaffected by these inhibitors. Furthermore, the cleavage of eIF4GI was observed in PV-infected MCF-7 cells lacking caspase-3. These data, and the fact that induction of apoptosis yields different eIF4GI cleavage fragments, indicate that caspases do not have a major role in the cleavage of eIF4GI during PV or FMDV infection.

scholarly journals Cleavage of Eukaryotic Translation Initiation Factor 4GII within Foot-and-Mouth Disease Virus-Infected Cells: Identification of the L-Protease Cleavage Site In Vitro

2004 ◽  
Vol 78 (7) ◽  
pp. 3271-3278 ◽  
Author(s):  
Alessandra Gradi ◽  
Nicole Foeger ◽  
Rebecca Strong ◽  
Yuri V. Svitkin ◽  
Nahum Sonenberg ◽  
...  
Keyword(s):  
Translation Initiation ◽  
Foot And Mouth Disease ◽  
Translation Initiation Factor ◽  
Initiation Factor ◽  
Eukaryotic Translation ◽  
Mouth Disease Virus ◽  
Eukaryotic Translation Initiation ◽  
Infected Cells ◽  
Mutant Forms

ABSTRACT Foot-and-mouth disease virus (FMDV) induces a very rapid inhibition of host cell protein synthesis within infected cells. This is accompanied by the cleavage of the eukaryotic translation initiation factor 4GI (eIF4GI). The cleavage of the related protein eIF4GII has now been analyzed. Within FMDV-infected cells, cleavage of eIF4GI and eIF4GII occurs with similar kinetics. Cleavage of eIF4GII is induced in cells and in cell extracts by the FMDV leader protease (Lpro) alone, generating cleavage products similar to those induced by enterovirus and rhinovirus 2A protease (2Apro). By the use of a fusion protein containing residues 445 to 744 of human eIF4GII, it was demonstrated that the FMDV Lpro specifically cleaves this protein between residues G700 and S701, immediately adjacent to the site (V699/G700) cleaved by rhinovirus 2Apro in vitro. The G700/S701 cleavage site does not correspond, by amino acid sequence alignment, to that cleaved in eIF4GI by the FMDV Lpro in vitro. Knowledge of the cleavage sites and the three-dimensional structures of the FMDV Lpro and rhinovirus 2Apro enabled mutant forms of the eIF4GII sequence to be generated that are differentially resistant to either one of these proteases. These results confirmed the specificity of each protease and showed that the mutant forms of the fusion protein substrate retained their correct sensitivity to other proteases.

scholarly journals Generation and genetic stability of tick-borne encephalitis virus mutants dependent on processing by the foot-and-mouth disease virus 3C protease

2012 ◽  
Vol 93 (3) ◽  
pp. 504-515 ◽  
Author(s):  
Sabrina Schrauf ◽  
Martina Kurz ◽  
Christian Taucher ◽  
Christian W. Mandl ◽  
Tim Skern
Keyword(s):  
Disease Virus ◽  
Cleavage Site ◽  
Protein C ◽  
Foot And Mouth Disease ◽  
Mouth Disease ◽  
3C Protease ◽  
C Terminus ◽  
Tick Borne Encephalitis ◽  
Mouth Disease Virus ◽  
Tick Borne Encephalitis Virus

Mature protein C of tick-borne encephalitis virus (TBEV) is cleaved from the polyprotein precursor by the viral NS2B/3 protease (NS2B/3pro). We showed previously that replacement of the NS2B/3pro cleavage site at the C terminus of protein C by the foot-and-mouth disease virus (FMDV) 2A StopGo sequence leads to the production of infectious virions. Here, we show that infectious virions can also be produced from a TBEV mutant bearing an inactivated 2A sequence through the expression of the FMDV 3C protease (3Cpro) either in cis or in trans (from a TBEV replicon). Cleavage at the C terminus of protein C depended on the catalytic activity of 3Cpro as well as on the presence of an optimized 3Cpro cleavage site. Passage of the TBEV mutants bearing a 3Cpro cleavage site either in the absence of 3Cpro or in the presence of a catalytically inactive 3Cpro led to the appearance of revertants in which protein C cleavage by NS2B/3pro had been regained. In three different revertants, a cleavage site for NS2B/3pro, namely RR*C, was now present, leading to an elongated protein C. Furthermore, two revertants acquired additional mutations in the C terminus of protein C, eliminating two basic residues. Although these latter mutants showed wild-type levels of early RNA synthesis, their foci were smaller and an accumulation of protein C in the cytoplasm was observed. These findings suggest a role of the positive charge of the C terminus of protein C for budding of the nucleocapsid and further support the notion that TBEV protein C is a multifunctional protein.

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