The Resistance Responses of Potato Plants to Potato Virus Y Are Associated with an Increased Cellular Methionine Content and an Altered SAM:SAH Methylation Index

Abstract

Plant-virus interactions are frequently influenced by elevated temperature, which often increases susceptibility to a virus, a scenario described for potato cultivar Chicago infected with potato virus Y (PVY). In contrast, other potato cultivars such as Gala may have similar resistances to PVY at both normal (22 °C) and high (28 °C) temperatures. To elucidate the mechanisms of temperature-independent antivirus resistance in potato, we analysed responses of Gala plants to PVY at different temperatures using proteomic, transcriptional and metabolic approaches. Here we show that in Gala, PVY infection generally upregulates the accumulation of major enzymes associated with the methionine cycle (MTC) independently of temperature, but that temperature (22 °C or 28 °C) may finely regulate what classes accumulate. The different sets of MTC-related enzymes that are up-regulated at 22 °C or 28 °C likely account for the significantly increased accumulation of S-adenosyl methionine (SAM), a key component of MTC which acts as a universal methyl donor in methylation reactions. In contrast to this, we found that in cultivar Chicago, SAM levels were significantly reduced which correlated with the enhanced susceptibility to PVY at high temperature. Collectively, these data suggest that MTC and its major transmethylation function determines resistance or susceptibility to PVY.

Keywords: isobaric tags for relative and absolute quantitation (iTRAQ); methionine cycle; plant virus resistance; potato virus Y.

Figure 1

A tentative model showing possible mechanisms determining susceptibility or resistance of potato plants to PVY. (A) Schematic representation of the methionine cycle (MTC) in healthy cells. Metabolite (shown in pink) flow directions are presented by arrows, while the enzymes involved in their bioconversion are marked in blue. CBL, cystathionine β-lyase; HCY, homocysteine; MET, methionine; MS, methionine synthase; THF, tetrahydrofolate; 5-MTHF, 5-methyltetrahydrofolate; 5,10-MTHF, 5,10-methylenetetrahydrofolate; MTHFR, methylene tetrahydrofolate reductase; SAH, S-adenosyl-homocysteine; SAHH, S-adenosyl-homocysteine hydrolase; SAM, S-adenosyl methionine; SAMS, S-adenosyl methionine synthetase; SHM, serine hydroxymethyltransferase; (B) Schematic representation of the changes in expression of key MTC-related enzymes and MTC metabolites in response to PVY. Black arrows show enzyme pathway and metabolite flow directions. Blue and red arrows indicate down- and up-regulation, respectively. PVY infection in Chicago plants at higher temperature causes down-regulation of key MTC (MS, SAMS, SAHH) and MTC-related folate cycle (SHM, MTHFR) enzymes, leading to the decrease of SAM and increase of SAH [5]. This would suppress SAM-dependent methylation reactions and decrease ethylene production, leading to a great increase in plant susceptibility to PVY. At normal temperature, PVY infection did not affect expression of MTC enzymes, and plants display much lower susceptibility to PVY compared with high temperature. PVY infection in Gala plants at higher temperature causes up-regulation of MS, SAMS and SHM, leading to the increase of SAM levels and SAM:SAH ratio (at 8 dpi). At higher temperature, Gala plants responded to PVY by up-regulation of CBL and SHM, also leading to increased SAM levels and SAM:SAH ratio. Gala plants remain resistant to PVY at both normal and high temperatures.

Figure 2

(A) Functional protein–protein association network of methionine cycle (MTC)-related differentially expressed proteins (DEPs) in PVY-infected Gala plants at normal and high temperature, constructed using STRING. Proteins are indicated with nodes, and links between proteins are represented by edges. CBL, cystathionine β-lyase; MS, methionine synthase; SAMS, S-adenosyl methionine synthetase; SHM, serine hydroxymethyltransferase; SAMDM, SAM dependent methyltransferase. (B) Heatmap showing the changes in abundance of key MTC related enzymes. PVY_22 °C _8dpi, infected Gala plants at 8 dpi at 22 °C; PVY_22 °C _14dpi, infected potato plants at 14 dpi at 22 °C; PVY_28 °C_8 dpi, infected potato plants at 8 dpi at 28 °C; PVY_28 °C _14 dpi, infected potato plants at 14 dpi at 28 °C.

Figure 3

Expression level of methionine cycle (MTC)-related protein genes using quantitative reverse transcription PCR [RT-qPCR]) in systemically infected leaves of Gala plants at 22 or 28 °C at five, seven, and eight days post-inoculation (dpi), as shown. CBL, cystathionine β-lyase; MS, methionine synthase; SAMS, S-adenosyl methionine synthetase; SHM, serine hydroxymethyltransferase; SAMDM, SAM dependent methyltransferase. Analysis of variance and Tukey’s HSD post hoc tests were performed for RT-qPCR data. *** p < 0.001.

Figure 4

Content of methionine cycle (MTC) metabolites and SAM:SAH ratios in systemically infected leaves of potato plants at 22 or 28 °C at 8-, 10-, and 14-days post-inoculation (dpi), as shown. Content of S-adenosylmethionine (SAM), S-adenosylhomocysteine (SAH), and homocysteine (HCY) was measured by ELISA. Content of methionine (MET) was measured using a methionine assay kit (fluorometric). Analysis of variance and Tukey’s HSD post hoc tests were performed for data. * p < 0.05; ** p < 0.01; *** p < 0.001.