Tethering of eIF4G to adenoviral mRNAs by viral 100k protein drives ribosome shunting

Abstract

Although most mRNAs initiate translation by 5 ribosome scanning, some small fraction of mammalian and viral mRNAs utilize either of two alternate mechanisms, known as internal ribosome entry and ribosome shunting. Ribosome shunting is a poorly understood form of initiation in which 40S ribosome subunits are loaded onto mRNA through interactions with the m 7 GTP cap, but then bypass large segments of the mRNA as directed by cis-acting RNA shunting elements and trans-acting protein factors. Here, we describe the molecular mechanism by which ribosome shunting occurs with high efficiency on adenovirus late mRNAs. We show that the viral 100k protein possesses a selective binding element for the 5 noncoding region (5 NCR) of viral late mRNAs (known as the tripartite leader), forms a complex with initiation factor eIF4G and poly(A)-binding protein (PABP), and strongly and selectively enhances the level of both factors and 40S ribosome subunits on viral mRNAs in polysomes. Mutational and biochemical studies demonstrate that the ability of 100k protein to bind both the tripartite leader and eIF4G are critical to promote a high level of ribosome shunting. A molecular mechanism for ribosome shunting is described by which enhanced binding of eIF4G and possibly PABP with 100k protein, and simultaneous interaction with the tripartite leader 5 NCR, drives 40S ribosome recruitment and initiation on mRNAs.

Inactivating intracellular antiviral responses during adenovirus infection

“…In this way, adenovirus compromises the ability of the eIF4F cap-binding complex to initiate translation on most mRNA. Adenovirus late mRNAs escape this block by using the modified eIF4F cap-binding complex to initiate translation by the unusual mechanism of ribosome shunting (Xi et al, 2004). Ribosome shunting allows adenovirus late mRNAs to escape the block to translation even though they are capped and polyadenylated in an identical manner to cellular mRNA (Schneider and Mohr, 2003).…”

“…The 100K protein accomplishes this as part of the modified eIF4F complex through its ability to bind specifically to late viral mRNA and the poly(A)-binding protein. The 100K protein contains both nonspecific (Adam and Dreyfuss, 1987;Riley and Flint, 1993) and sequence-specific RNAbinding domains (Xi et al, 2004(Xi et al, , 2005. As part of eIF4F, the 100K protein binds the tripartite leader in a tyrosine phosphorylation-specific manner (Xi et al, 2004(Xi et al, , 2005.…”

“…The 100K protein contains both nonspecific (Adam and Dreyfuss, 1987;Riley and Flint, 1993) and sequence-specific RNAbinding domains (Xi et al, 2004(Xi et al, , 2005. As part of eIF4F, the 100K protein binds the tripartite leader in a tyrosine phosphorylation-specific manner (Xi et al, 2004(Xi et al, , 2005. Therefore, through interactions with viral mRNA, eIF4G and the poly(A)-binding protein, the 100K protein recruits factors required for translation of viral mRNA in the face of the overall inhibition to cap-dependent translation.…”

To replicate or not to replicate: achieving selective oncolytic virus replication in cancer cells through translational control

“…Following recruitment of the 40S ribosome to the 5 0 end through an association with eIF4F and eIF3, the search for the AUG codon initially proceeds through a scanning mechanism; however, once the ribosome encounters the tripartite leader, it appears to be transferred to the AUG by a nonlinear translocation process (Yueh and Schneider, 1996). The mechanism through which 100 K promotes shunting involves both its binding to mRNA, which is regulated by tyrosine phosphorylation, and eIF4G (Xi et al, 2004(Xi et al, , 2005. The interaction of eIF4G with 100 K displaces the mnk kinase, resulting in the accumulation of unphosphorylated eIF4E.…”

“…While the resulting block to replication in normal cells likely results from the combined effects of retaining multiple late mRNAs that specify the various components of the adenovirus capsid within nuclei, it is particularly noteworthy that the late mRNA encoding the 100 K protein is among this class of sequestered transcripts. One of the functions of 100 K is to promote the translation of adenoviral late mRNAs, all of which have a common tripartite leader sequence spliced onto their capped 5 0 end (Cuesta et al, 2000;Xi et al, 2004). The tripartite leader is in fact a cis-acting genetic element that promotes translational initiation by ribosome shunting (Yueh and Schneider, 1996).…”