Structural characterization of the Z RING-eIF4E complex reveals a distinct mode of control for eIF4E

Volpon, Laurent; Osborne, Michael J.; Capul, Althea A.; Torre, Juan Carlos de la; Borden, Katherine L. B.

doi:10.1073/pnas.0909877107

Cited by 67 publications

(73 citation statements)

References 41 publications

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“…S4 A and B). However, eIF4E interaction was only detectable by immunoblot, consistent with previously published results (12), and eIF4E alone was not sufficient to affect Z-L complex formation or Z-induced regulation of viral RNA synthesis (Fig. S4C).…”

Section: Resultssupporting

confidence: 79%

“…The Z gene is highly conserved across all arenavirus species and encodes a protein organized as an unstructured amino and carboxyl terminus bracketing a really interesting new gene (RING) domain that coordinates a pair of structural zinc ions (12). Z has been implicated in many facets of the arenavirus replication cycle and is the primary driving force of virion maturation (13).…”

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confidence: 99%

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Arenavirus Z protein controls viral RNA synthesis by locking a polymerase–promoter complex

Kranzusch

Whelan

2011

Proc. Natl. Acad. Sci. U.S.A.

103

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Arenaviruses form a noncytolytic infection in their rodent hosts, yet can elicit severe hemorrhagic disease in humans. How arenaviruses regulate gene expression remains unclear, and further understanding may provide insight into the dichotomy of these disparate infection processes. Here we reconstitute arenavirus RNA synthesis initiation and gene expression regulation in vitro using purified components and demonstrate a direct role of the viral Z protein in controlling RNA synthesis. Our data reveal that Z forms a species-specific complex with the viral polymerase (L) and inhibits RNA synthesis initiation by impairing L catalytic activity. This Z-L complex locks the viral polymerase in a promoter-bound, catalytically inactive state and may additionally ensure polymerase packaging during virion maturation. Z modulates host factors involved in cellular translation, proliferation, and antiviral signaling. Our data defines an additional role in governing viral RNA synthesis, revealing Z as the center of a network of host and viral connections that regulates viral gene expression.transcription factor | Machupo virus | matrix protein | negative-strand RNA virus | gene regulation T ranscription initiation is a fundamental focal point of gene expression regulation in all orders of life. Modulation of RNA synthesis affords an important response to stress, alterations in growth conditions, and environmental cues. Although initiation of RNA synthesis is often indirectly controlled through gene promoters and accessory signaling elements, regulation also occurs through factors directly interacting with and altering the catalytic potential of the RNA synthesis machinery itself (1). Studies with phage T7 and the T7 lysozyme repressor system have provided an important paradigm for control of viral RNA transcription (2, 3), but these processes remain a mystery for many eukaryotic viruses and medically relevant pathogens. Gene expression regulation is particularly critical for negative-sense RNA viruses of the family Arenaviridae, which must respond to input from the host environment to maintain a noncytolytic infection in their rodent reservoir (4). In stark contrast to this persistent infection, arenavirus infection in humans can lead to severe hemorrhagic fever disease and many arenaviruses represent important emerging pathogens (5). Intriguingly, recent evidence indicates that the balance between persistent and acute viral infection can be altered by a single mutation within the viral polymerase (6). Mechanistic insight into how arenaviruses tune gene expression and control viral RNA synthesis is required to further our understanding of the response of arenavirus infection to various host and cellular environments.Arenaviruses are the simplest of all hemorrhagic fever viruses, with only four viral proteins. The viral genome consists of two segments of single-stranded negative-sense RNA, where each segment encodes two proteins in an ambisense orientation. The small segment encodes for the viral glycoprotein, essential for entr...

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Section: Resultssupporting

confidence: 79%

mentioning

confidence: 99%

Arenavirus Z protein controls viral RNA synthesis by locking a polymerase–promoter complex

Kranzusch

Whelan

2011

Proc. Natl. Acad. Sci. U.S.A.

103

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“…The Z protein of arenaviruses contains a conserved 37-aminoacid RING domain that coordinates two zinc ions and that is surrounded by less conserved N-and C-terminal regions (22,51). Several lines of experimental evidence indicate that the Z protein has multiple functions during the viral life cycle.…”

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confidence: 99%

Molecular Determinants of Arenavirus Z Protein Homo-Oligomerization and L Polymerase Binding

Loureiro

Wilda

Macleod

et al. 2011

J Virol

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The arenavirus Z is a zinc-binding RING protein that has been implicated in multiple functions during the viral life cycle. These roles of Z involve interactions with viral and cellular proteins that remain incompletely understood. In this regard, Z inhibits viral RNA transcription and replication through direct interaction with the viral L polymerase. Here, we defined the L-binding domain of Tacaribe virus (TCRV) Z protein and the structural requirements mediating Z homo-oligomerization. By using site-directed mutagenesis, coimmunoprecipitation, and functional assays, we showed that residues R37, N39, W44, L50, and Y57, located around the zinc coordination site I, play a critical role in the Z-L interaction. We also found that Z protein from either TCRV or the pathogenic Junin virus (JUNV) self-associates into oligomeric forms in mammalian cells. Importantly, mutation of the myristoylation site, the strictly conserved residue G at position 2, severely impaired the ability of both TCRV Z and JUNV Z to self-interact as well as their capacity to accumulate at the plasma membrane, strongly suggesting that Z homo-oligomerization is associated with its myristoylation and cell membrane targeting. In contrast, disruption of the RING structure or substitution of W44 or N39, which are critical for L protein recognition, did not affect Z self-binding. Overall, the data presented here indicate that homo-oligomerization is not a requirement for Z-L interaction or Z-mediated polymerase activity inhibition.

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“…Detailed structural information is available on eIF4E from different species, in complex with cap analogs and with peptides derived from eIF4G and 4E-BPs. These peptides bind in a similar mode to a conserved surface on eIF4E, opposite and distal to the m 7 G cap-binding pocket (Marcotrigiano et al 1997(Marcotrigiano et al , 1999Matsuo et al 1997;Niedzwiecka et al 2002a;Gross et al 2003;Tomoo et al 2005;Volpon et al 2006Volpon et al , 2010Brown et al 2007Brown et al , 2009Monzingo et al 2007;Rosettani et al 2007;Ashby et al 2011;Liu et al 2011;Siddiqui et al 2012) (pdb id. : 3AM7).…”

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confidence: 99%

Crystal structure of a minimal eIF4E–Cup complex reveals a general mechanism of eIF4E regulation in translational repression

Kinkelin

Veith

Grünwald

et al. 2012

RNA

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Cup is an eIF4E-binding protein (4E-BP) that plays a central role in translational regulation of localized mRNAs during early Drosophila development. In particular, Cup is required for repressing translation of the maternally contributed oskar, nanos, and gurken mRNAs, all of which are essential for embryonic body axis determination. Here, we present the 2.8 Å resolution crystal structure of a minimal eIF4E-Cup assembly, consisting of the interacting regions of the two proteins. In the structure, two separate segments of Cup contact two orthogonal faces of eIF4E. The eIF4E-binding consensus motif of Cup (YXXXXLF) binds the convex side of eIF4E similarly to the consensus of other eIF4E-binding proteins, such as 4E-BPs and eIF4G. The second, noncanonical, eIF4E-binding site of Cup binds laterally and perpendicularly to the eIF4E b-sheet. Mutations of Cup at this binding site were shown to reduce binding to eIF4E and to promote the destabilization of the associated mRNA. Comparison with the binding mode of eIF4G to eIF4E suggests that Cup and eIF4G binding would be mutually exclusive at both binding sites. This shows how a common molecular surface of eIF4E might recognize different proteins acting at different times in the same pathway. The structure provides insight into the mechanism by which Cup disrupts eIF4E-eIF4G interaction and has broader implications for understanding the role of 4E-BPs in translational regulation.

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Structural characterization of the Z RING-eIF4E complex reveals a distinct mode of control for eIF4E

Cited by 67 publications

References 41 publications

Arenavirus Z protein controls viral RNA synthesis by locking a polymerase–promoter complex

Arenavirus Z protein controls viral RNA synthesis by locking a polymerase–promoter complex

Molecular Determinants of Arenavirus Z Protein Homo-Oligomerization and L Polymerase Binding

Crystal structure of a minimal eIF4E–Cup complex reveals a general mechanism of eIF4E regulation in translational repression

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