Abstract
Cassava brown streak disease (CBSD), dubbed the "Ebola of plants", is a serious threat to food security in Africa caused by two viruses of the family Potyviridae: cassava brown streak virus (CBSV) and Ugandan (U)CBSV. Intriguingly, U/CBSV, along with another member of this family and one secoviridae, are the only known RNA viruses encoding a protein of the Maf/ham1-like family, a group of widespread pyrophosphatase of non-canonical nucleotides (ITPase) expressed by all living organisms. Despite the socio-economic impact of CDSD, the relevance and role of this atypical viral factor has not been yet established. Here, using an infectious cDNA clone and reverse genetics, we demonstrate that UCBSV requires the ITPase activity for infectivity in cassava, but not in the model plant Nicotiana benthamiana. HPLC-MS/MS experiments showed that, quite likely, this host-specific constraint is due to an unexpected high concentration of non-canonical nucleotides in cassava. Finally, protein analyses and experimental evolution of mutant viruses indicated that keeping a fraction of the yielded UCBSV ITPase covalently bound to the viral RNA-dependent RNA polymerase (RdRP) optimizes viral fitness, and this seems to be a feature shared by the other members of the Potyviridae family expressing Maf/ham1-like proteins. All in all, our work (i) reveals that the over-accumulation of non-canonical nucleotides in the host might have a key role in antiviral defense, and (ii) provides the first example of an RdRP-ITPase partnership, reinforcing the idea that RNA viruses are incredibly versatile at adaptation to different host setups.
Highlights
The family Potyviridae is the largest and most socio-economically relevant group of plantinfecting RNA viruses
We found that HAM1 is required for Ugandan cassava brown streak virus (UCBSV) to infect cassava, where its pyrophosphatase activity resulted critical, but not to propagate in the model plant Nicotiana benthamiana
Unexpected high levels of inosine triphosphate (ITP)/XTP non-canonical nucleotides found in cassava, and the known flexibility of RNA viruses to incorporate additional factors when required, supports the idea that the high concentration of ITP/XTP worked as a selection pressure to promote the acquisition of HAM1 into the virus in order to promote a successful infection
Summary
The family Potyviridae is the largest and most socio-economically relevant group of plantinfecting RNA viruses. Potyvirids (members of the family Potyviridae) share common features, such as (i) monopartite (except for a few bipartite viruses) and positive sense single-stranded RNA (+ssRNA) genome, (ii) transmission mediated by vectors, and (iii) picorna-like gene expression strategy based on large polyproteins further processed by viral-encoded proteinases [1–3]. Of relevance to this study, NIaPro is a cis- and trans-acting proteinase that releases most of the mature factors from the polyprotein [4,5], and NIb is a RNA-dependent RNA polymerase (RdRP) that replicates the viral genome [6,7]. With seven members described so far, the Ipomovirus genus is the most versatile group of potyvirids in term of genome organization, since only two of them follow the most common arrangement mentioned above [8]. CBSD is considered among the seven most detrimental plant diseases in the world for its impact on the economy and food security in Africa, where it causes about 750 million US$ annual losses just in Tanzania, Uganda, Kenya and Malawi [14,15]
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