Formation of large viroplasms and virulence of Cauliflower mosaic virus in turnip plants depend on the N-terminal EKI sequence of viral protein TAV

Cauliflower mosaic virus (CaMV) TAV protein (TransActivator/Viroplasmin) plays a pivotal
role during the infection cycle since it activates translation reinitiation of viral polycistronic
RNAs and suppresses RNA silencing. It is also the major component of cytoplasmic electron-dense inclusion bodies (EDIBs) called viroplasms that are particularly evident in cells
infected by the virulent CaMV Cabb B-JI isolate. These EDIBs are considered as virion factories, vehicles for CaMV intracellular movement and reservoirs for CaMV transmission by
aphids. In this study, focused on different TAV mutants in vivo, we demonstrate that three
physically separated domains collectively participate to the formation of large EDIBs: the Nterminal EKI motif, a sequence of the MAV domain involved in translation reinitiation and a
C-terminal region encompassing the zinc finger. Surprisingly, EKI mutant TAVm3, corresponding to a substitution of the EKI motif at amino acids 11–13 by three alanines (AAA),
which completely abolished the formation of large viroplasms, was not lethal for CaMV but
highly reduced its virulence without affecting the rate of systemic infection. Expression of
TAVm3 in a viral context led to formation of small irregularly shaped inclusion bodies, mild
symptoms and low levels of viral DNA and particles accumulation, despite the production of
significant amounts of mature capsid proteins. Unexpectedly, for CaMV-TAVm3 the formation of viral P2-containing electron-light inclusion body (ELIB), which is essential for CaMV
aphid transmission, was also altered, thus suggesting an indirect role of the EKI tripeptide in
CaMV plant-to-plant propagation. This important functional contribution of the EKI motif in
CaMV biology can explain the strict conservation of this motif in the TAV sequences of all
CaMV isolates.