2b Protein is essential to induce a novel gradual cell death in Zucchini yellow mosaic virus-inoculated cucumber cotyledon co-infected with Cucumber mosaic virus

Cucumber cotyledons inoculated with Cucumber mosaic virus (CMV, Pepo strain) or Zucchini yellow mosaic virus (ZYMV, Z5-1 isolate) developed either mild chlorotic spots or no symptoms. Cotyledons treated with CMV plus ZYMV also developed mild chlorotic spots. However, plants ZYMV-inoculated cotyledons had veinal yellowing and gradual cell death by 20 days postinoculation (dpi) when co-inoculated with CMV on the other cotyledon. When analyzing this synergism, an enzyme-linked immunosorbent assay showed that CMV gradually increased in CMV-inoculated cotyledons of plants, with the other cotyledon mock- or ZYMV-inoculated. However, CMV significantly increased at 9 to 14 dpi in the ZYMV-inoculated cotyledons of plants co-infected with CMV. ZYMV similarly increased in cotyledon pairs of both co-infected and singly infected plants. Inoculation with Pepodelta2b, a modified Pepo-CMV that lacks translation of the 2b protein, revealed that Pepodelta2b without the 2b protein systemically infected cucumber but induced no symptoms on cotyledons or true leaves. Plants with a ZYMV-inoculated cotyledon and co-infected with Pepodelta2b did not undergo cell death; nevertheless, Pepodelta2b was at high levels comparable to levels of CMV in the ZYMV-inoculated cotyledon. The 2b protein thus seems essential for induction of the novel gradual cell death in ZYMV-inoculated cotyledons of cucumbers co-infected with CMV.[PUBLICATION ABSTRACT]