Foxtail mosaic virus: A Viral Vector for Protein Expression in Cereals

Abstract

Rapid and cost-effective virus-derived transient expression systems for plants are invaluable in elucidating gene function and are particularly useful in plant species for which transformation-based methods are unavailable or are too time and labor demanding, such as wheat () and maize (). The virus-mediated overexpression (VOX) vectors based on and described previously for these species are incapable of expressing free recombinant proteins of more than 150 to 250 amino acids, are not suited for high-throughput screens, and have other limitations. In this study, we report the development of a VOX vector based on a monopartite single-stranded positive sense RNA virus, (genus). In this vector, PV101, the gene of interest was inserted downstream of the duplicated subgenomic promoter of the viral coat protein gene, and the corresponding protein was expressed in its free form. The vector allowed the expression of a 239-amino acid-long GFP in both virus-inoculated and upper uninoculated (systemic) leaves of wheat and maize and directed the systemic expression of a larger approximately 600-amino acid protein, GUSPlus, in maize. Moreover, we demonstrated that PV101 can be used for in planta expression and functional analysis of apoplastic pathogen effector proteins such as the host-specific toxin ToxA of Therefore, this VOX vector opens possibilities for functional genomics studies in two important cereal crops.

Applying the latest advances in genomics and phenomics for trait discovery in polyploid wheat

“…The use of virus‐induced gene silencing (VIGS) in wheat and other cereals has been highly successful in characterizing the impact of various genes involved in abiotic and biotic stress response (Cakir et al ., ; Ramegowda et al ., ; Lee et al ., ). These VIGS systems are now well established for use in wheat, and more recently advances in virus‐mediated overexpression have demonstrated the feasibility of their use in wheat for both small and large proteins (Bouton et al ., ; Cheuk and Houde, ). Beyond viral expression systems, other possibilities for transient expression in wheat exist.…”

Harnessed viruses in the age of metagenomics and synthetic biology: an update on infectious clone assembly and biotechnologies of plant viruses

“…Current chemical DNA synthesis prices are not compatible with routine cloning, although a sharp price drop catalysed by new technology developments is expected in the near future (Schindler et al, 2018). In the plant virology field, infectious clones have been reported for delivery of tobamovirus, tombusvirus and potexvirus genomes synthesized entirely from scratch (Bouton et al, 2018;Cooper, 2014;Lovato et al, 2014). Due to errors in the reference sequence used, a first version of the synthetic tobamovirus genome was not infectious.…”

Protein expression and gene editing in monocots using foxtail mosaic virus vectors

“…Inverted‐repeats carried at this position were most efficient at inducing VIGS (Liu et al, ). Bouton et al () also duplicated the CP promoter and demonstrated that FoMV could be used to transiently express marker and fungal effector proteins from this position in N. benthamiana , wheat, and maize. Disarmed FoMV vectors for transient gene expression have also been developed by substitution of the TGB or CP with genes of interest.…”

“…The development of viral vectors for monocot plants has been emerging rapidly in recent years (Lee, Hammond-Kosack, & Kanyuka, 2015). To date, at least eight different viruses have been developed into viral vectors for monocots, including barley stripe mosaic virus (BSMV) (Lee, Hammond-Kosack, & Kanyuka, 2012;Scofield, Huang, Brandt, & Gill, 2005), brome mosaic virus (BMV) (Ding, Schneider, Chaluvadi, Mian, & Nelson, 2006), cymbidium mosaic virus (CymMV) (Hsieh et al, 2013), rice tungro bacilliform virus (RTBV) (Purkayastha, Mathur, Verma, Sharma, & Dasgupta, 2010), wheat streak mosaic virus (WSMV) (Choi, Stenger, Morris, & French, 2000;Tatineni, McMechan, Hein, & French, 2011), bamboo mosaic virus (BaMV) together with its associated satellite RNA (Liou, Huang, Hu, Lin, & Hsu, 2014), cucumber mosaic virus (CMV) (Wang et al, 2016), barley yellow striate mosaic virus (BYSMV) (Gao et al, 2019), and foxtail mosaic virus (FoMV) (Bouton et al, 2018;Liu et al, 2016;Mei & Whitham, 2018;Mei, Zhang, Kernodle, Hill, & Whitham, 2016). Seven of the virus vectors are designed for VIGS applications (BSMV, BMV, CymMV, RTBV, BaMV, CMV, and FoMV), and four can be used for systemic gene expression (BSMV, BYSMV, FoMV, and WSMV).…”

“…Previous work by us and others demonstrated that FoMV is capable of systemic infection and inducing VIGS in maize Mei & Whitham, 2018;Mei et al, 2016) or expressing proteins (Bouton et al, 2018). FoMV is a member of the genus Potexvirus, which has a single-stranded, positive-sense genomic RNA.…”