1.11.22
A SINGLE AMINO ACID IN COAT PROTEIN OF CUCUMBER MOSAIC VIRUS CONTROLS TWO DISTINCT HOST RESPONSES IN VIRUS-INOCULATED LEAVES OF ARABIDOPSIS THALIANA AND NICOTIANA TABACUM

H TAKAHASHI1, T SHIGYO1, M SUZUKI2, T TERAOKA1, Y EHARA3 and D HOSOKAWA1

1Tokyo University of Agriculture and Technology, Saiwai-cho, Fuchu, Tokyo 183-8509, Japan; 2 Japan Tobacco Inc., Umegaoka, Aoba-ku, Yokohama 227, Japan; 3 Tohoku University, Tsutsumidori-Amamiyamachi, Aoba-ku, Sendai 981, Japan

Background and objectives
Cucumber mosaic virus (CMV) is one of the best studied viruses and includes many strains which infect various plant species and show different symptoms. A yellow strain of CMV [CMV(Y)] induces severe chlorotic spot symptoms in virus-inoculated leaves of systemically infected tobacco (Nicotiana tabacum cv. Ky57), although an ordinary strain of CMV [CMV(O)] does not show any symptoms on the inoculated leaves. Severe chlorotic spot symptoms in CMV(Y)-inoculated Ky57 leaves was controlled by the interaction between CMV coat protein gene and putative barley genes (yb1 and yb2) of Ky57, although the amino acid position(s) in the coat protein, which determined the expression of severe chlorotic spot symptoms, has not been defined [1]. On the other hand, CMV(Y) induced the HR in Arabidopsis thaliana ecotype C24 but not in ecotype Columbia, although CMV(O) systemically infected both ecotypes. The HR in CMV(Y)-inoculated C24 was controlled by the interaction between CMV RNA3 and a single dominant resistance gene, RCY1, of C24 [2]. In this report, the virus gene on RNA3, which controls the HR in C24 and severe chlorotic spot formation in Ky57, was characterized.

Results and conclusions
To define the determinant in RNA3 for the distinct host responses in C24 and Ky57, the chimeric cDNAs of RNA3 between CMV(Y) and CMV(O) were constructed, and RNA3 in vitro transcribed from the cDNA, was inoculated to C24 and Ky57 with CMV(Y) RNA1+2. The infectious RNA3, in which CMV(O) virus movement protein and CMV(Y) coat protein (OMP/YCP) were encoded, induced the HR in C24 and severe chlorotic spot symptoms in Ky57, although the infectious RNA3 (YMP/OCP) systemically infected both host plants and sometimes showed very mild chlorosis on the inoculated leaves of Ky57. The result indicated that CMV(Y) coat protein gene acts as the main determinant in both the HR of CMV(Y)-inoculated C24 and severe chlorotic spot formation on CMV(Y)-inoculated Ky57. For further analysis of the domain in CMV(Y) coat protein which determines the two distinct host responses, a series of chimeric infectious RNA3 were constructed by exchanging restriction fragments in coat protein gene between CMV(Y) and CMV(O) at the cDNA level. The chimeric infectious RNA3s exchanged NruI-XhoI fragment in coat protein gene at the cDNA level, changed the responses to CMV in both C24 and Ky57. In comparison of the amino acid sequence corresponding to the NruI-XhoI fragment between CMV(Y) and CMV(O), only Ser at the 129 amino acid position in CMV(Y) was different from Pro in CMV(O). Infectious RNA3, in which only Ser at 129 amino acid position in CMV(Y) coat protein was substituted for Pro, did not induce either the HR and severe chlorotic spot symptoms on the inoculated leaves of C24 and Ky57. These results indicated that a single amino acid at 129 position in coat protein determined the two distinct host responses; the HR to CMV in Arabidopsis thaliana ecotype C24 and severe chlorotic spot symptom formation in the inoculated leaves of CMV systemically-infected Nicotiana tabacum cv. Ky57.

References
1. Takahashi H, Ehara Y, 1993. Mol. Plant-Microbe Interact. 6, 182-189.
2. Takahashi H et al., 1994. Plant J. 6, 369-377.