Foxtail mosaic virus: a new viral vector for protein expression in cereals

Rapid and cost-effective virus-derived transient expression systems for plants are invaluable in elucidating gene function. These are particularly useful in the case of plant species for which transformation-based methods are either not yet developed, or are too time- and labor-demanding, such as wheat and maize. The Virus-mediated overexpression (VOX) vectors based on Barley stripe mosaic virus (BSMV) or Wheat streak mosaic virus (WSMV) previously described for these species are incapable of expressing free recombinant proteins >150-250 amino-acids (aa), not suited for high throughput screens, and have other limitations. In this study, we report the development of a new VOX vector based on a monopartite single-stranded positive sense RNA virus, Foxtail mosaic virus (FoMV, genus Potexvirus). The gene of interest is inserted downstream of a duplicated sub-genomic promoter of the viral coat protein gene and the corresponding protein is expressed in its free form. This new vector, PV101, allowed expression of a 239 aa-long green fluorescent protein (GFP) in both virus inoculated and upper uninoculated (systemic) leaves of wheat and maize, and directed systemic expression of a larger ca. 600 aa 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 host-specific toxin ToxA of Parastagonospora nodorum. Therefore, this new VOX vector opens new possibilities for functional genomics studies in two of the most important cereal crops

Transcriptomic Characterization of a Synergistic Genetic Interaction during Carpel Margin Meristem Development in Arabidopsis thaliana

In flowering plants the gynoecium is the female reproductive structure. In Arabidopsis thaliana ovules initiate within the developing gynoecium from meristematic tissue located along the margins of the floral carpels. When fertilized the ovules will develop into seeds. SEUSS (SEU) and AINTEGUMENTA (ANT) encode transcriptional regulators that are critical for the proper formation of ovules from the carpel margin meristem (CMM). The synergistic loss of ovule initiation observed in the seu ant double mutant suggests that SEU and ANT share overlapping functions during CMM development. However the molecular mechanism underlying this synergistic interaction is unknown. Using the ATH1 transcriptomics platform we identified transcripts that were differentially expressed in seu ant double mutant relative to wild type and single mutant gynoecia. In particular we sought to identify transcripts whose expression was dependent on the coordinated activities of the SEU and ANT gene products. Our analysis identifies a diverse set of transcripts that display altered expression in the seu ant double mutant tissues. The analysis of overrepresented Gene Ontology classifications suggests a preponderance of transcriptional regulators including multiple members of the REPRODUCTIVE MERISTEMS (REM) and GROWTH-REGULATING FACTOR (GRF) families are mis-regulated in the seu ant gynoecia. Our in situ hybridization analyses indicate that many of these genes are preferentially expressed within the developing CMM. This study is the first step toward a detailed description of the transcriptional regulatory hierarchies that control the development of the CMM and ovule initiation. Understanding the regulatory hierarchy controlled by SEU and ANT will clarify the molecular mechanism of the functional redundancy of these two genes and illuminate the developmental and molecular events required for CMM development and ovule initiation