Up-regulation ofLSB1/GDU3affects geminivirus infection by activating the salicylic acid pathway

Abstract

SUMMARYGeminiviruses include a large number of single-stranded DNA viruses that are emerging as useful tools to dissect many fundamental processes in plant hosts. However, there have been no reports yet regarding the genetic dissection of the geminivirus-plant interaction. Here, a high-throughput approach was developed to screen Arabidopsis activation-tagged mutants which are resistant to geminivirus Beet severe curly top virus (BSCTV) infection. A mutant, lsb1 (less susceptible to BSCTV 1), was identified, in which BSCTV replication was impaired and BSCTV infectivity was reduced. We found that the three genes closest to the T-DNA were up-regulated in lsb1, and the phenotypes of lsb1 could only be recapitulated by the overexpression of GDU3 (GLUTAMINE DUMPER 3), a gene implicated in amino acid transport. We further demonstrated that activation of LSB1/GDU3 increased the expression of components in the salicylic acid (SA) pathway, which is known to counter geminivirus infection, including the upstream regulator ACD6. These data indicate that up-regulation of LSB1/GDU3 affects BSCTV infection by activating the SA pathway. This study thus provides a new approach to study of the geminivirus-host interaction.

BSCTV C2 Attenuates the Degradation of SAMDC1 to Suppress DNA Methylation-Mediated Gene Silencing in Arabidopsis

“…Thus, we hypothesized that changes affecting host DNA methylation may also affect host antiviral defenses and geminivirus infectivity. To test this idea, we first compared the infectivity of c2 -mutant BSCTV to that of intact BSCTV using an A. tumefaciens spraying method described previously (Chen et al, 2010). The c2 -mutant BSCTV showed milder and delayed symptoms, such as longer inflorescence, mild curling inflorescence, and more production of lateral branches ( Figure 5A).…”

“…SD medium minus Leu and Trp ensure the maintenance of both expression plasmids. The interactions were confirmed by assessing b-galactosidase activity using a filter-lift assay (Breeden and Nasmyth, 1985) The construct containing full-length BSCTV C2 was generated by PCR using pCambia1300-BSCTV (1.8 copy) (Chen et al, 2010). Constructs containing defined regions of SAMDC1 were generated by different restriction endonuclease digestions of full-length SAMDC1 cDNA: truncation from amino acids 1 to 83 was produced by KpnI; truncation from amino acids 182 to 292 was produced by EcoRI and XhoI; truncation from amino acids 292 to 366 was produced by XhoI; and truncation from amino acids 226 to 366 was produced by EcoRI.…”

“…Rosette leaves of 4-week-old plants were agroinoculated with BSCTV using an established A. tumefaciens spraying method described previously (Chen et al, 2010). The concentration of the suspension of A. tumefaciens strain EHA105 (Hood et al, 1993) carrying pCambia1300-BSCTV (1.8 copy) or the C2 mutant was at a dose of OD 600 = 2.0.…”

“…Total DNA was extracted using a CTAB buffer as described (Chen et al, 2010). Genomic DNA was stained using ethidium bromide as a loading control.…”

“…Plants have evolved various mechanisms to respond to viral invasion, such as pathways mediated by gene silencing, salicylic acid, and metabolism regulation (Herbers et al, 1996;Vanitharani et al, 2005;Bisaro, 2006;Ascencio-Ibá ñ ez et al, 2008;Chen et al, 2010). Among them, gene silencing-mediated antiviral defense is a wellconserved and effective pathway.…”