Jun Ohnishi

Amino acid substitution in the coat protein of Melon necrotic spot virus causes loss of binding to the surface of Olpidium bornovanus zoospores

Melon necrotic spot virus (MNSV) is transmitted by the fungus Olpidiumbornovanus. In this study, we used immunofluorescence microscopy to detect MNSV particles over the entire surface of the O. bornovanus zoospore; MNSV particles were not detected on the related fungus O. virulentus, which cannot transmit MNSV. The amino acid substitution Ile → Phe at position 300 in the MNSV coat protein resulted in loss of both specific binding and fungal transmission, while virion assembly and biological aspects were unaffected. Taken together, these results suggest that the MNSV coat protein acts as a ligand to the O. bornovanus zoospore as part of a fungal-vector transmission system.

Structure and enzyme expression in photosynthetic organs of the atypical C4 grass Arundinella hirta

In its leaf blade, Arundinella hirta has unusual Kranz cells that lie distant from the veins (distinctive cells; DCs), in addition to the usual Kranz units composed of concentric layers of mesophyll cells (MCs) and bundle sheath cells (BSCs; usual Kranz cells) surrounding the veins. We examined whether chlorophyllous organs other than leaf blades—namely, the leaf sheath, stem, scale leaf, and constituents of the spike—also have this unique anatomy and the C4 pattern of expression of photosynthetic enzymes. All the organs developed DCs to varying degrees, as well as BSCs. The stem, rachilla, and pedicel had C4-type anatomy with frequent occurrence of DCs, as in the leaf blade. The leaf sheath, glume, and scale leaf had a modified C4 anatomy with MCs more than two cells distant from the Kranz cells; DCs were relatively rare. An immunocytochemical study of C3 and C4 enzymes revealed that all the organs exhibited essentially the same C4 pattern of expression as in the leaf blade. In the scale leaf, however, intense expression of ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco) occurred in the MCs as well as in the BSCs and DCs. In the leaf sheath, the distant MCs also expressed Rubisco. In Arundinella hirta, it seems that the ratio of MC to Kranz cell volumes, and the distance from the Kranz cells, but not from the veins, affects the cellular expression of photosynthetic enzymes. We suggest that the main role of DCs is to keep a constant quantitative balance between the MCs and Kranz cells, which is a prerequisite for effective C4 pathway operation.

Co-transmission of Tomato yellow leaf curl virus (TYLCV)-Mld and TYLCV-IL by the whitefly Bemisia tabaci

The ability of the whitefly Bemisia tabaci to transmit two strains of Tomato yellow leaf curl virus, the Israel and Mild strains, was studied after serial transfers of individual whiteflies that were viruliferous for both strains to tomato plants. After single whiteflies had successive acquisition feedings first on a single plant infected with one strain and then on a plant infected with the other strain, the single whiteflies later transmitted intermittently one, the other, or both strains to the test plants during serial transfers at 1-day intervals. Because both strains were found in the head, abdomen, and legs dissected from whiteflies during the retention period after the two successive acquisition feedings, both strains apparently circulate from midgut cells to salivary glands through the hemolymph.

Analysis of the Mild strain of tomato yellow leaf curl virus, which overcomes Ty-2 gene–mediated resistance in tomato line H24

In tomato line H24, an isolate of the Mild (Mld) strain of tomato yellow leaf curl virus (TYLCV-Mld [JR:Kis]) overcomes Ty-2 gene–mediated resistance and causes typical symptoms of tomato yellow leaf curl disease (TYLCD). No systemic infection with visible symptoms or accumulation of viral DNA in the upper leaves was observed in H24 challenged with another isolate, TYLCV-IL (TYLCV-IL [JR:Osaka]), confirming that H24 is resistant to the IL strain. To elucidate the genomic regions that cause the breakdown of the Ty-2 gene–mediated resistance, we constructed a series of chimeras by swapping genes between the two strains. A chimeric virus that had the overlapping C4/Rep region of the Mld strain in the context of the IL strain genome, caused severe TYLCD in H24 plants, suggesting that the overlapping C4/Rep region of the Mld strain is associated with the ability of this strain to overcome Ty-2 gene–mediated resistance.

Structure and immunocytochemical localization of photosynthetic enzymes in the lamina joint and sheath pulvinus of the C4 grass Arundinella hirta

The C4 grass Arundinella hirta exhibits a unique C4 anatomy, with isolated Kranz cells (distinctive cells) and C4-type expression of photosynthetic enzymes in the leaf sheath and stem as well as in the leaf blade. The border zones between these organs are pale green. Those between the leaf blade and sheath and between the sheath and stem are called the lamina joint and sheath pulvinus, respectively, and are involved in gravity sensing. We investigated the structure and localization of C3 and C4 photosynthetic enzymes in these tissues. In both zones the epidermis lacked stomata. The inner tissue was composed of parenchyma cells and vascular bundles. The parenchyma cells were densely packed with small intercellular spaces and contained granal chloroplasts with large starch grains. No C4-type cellular differentiation was recognized. Western blot analysis showed that the lamina joint and pulvinus accumulated substantial amounts of phosphoenolpyruvate carboxylase (PEPC), pyruvate,Pi dikinase (PPDK), and ribulose 1,5-bisphosphate carboxylase/oxygenase (rubisco). Immunogold electron microscopy revealed PEPC in the cytosol and both PPDK and rubisco in the chloroplasts of parenchyma cells, suggesting the occurrence of C3 and C4 enzymes within a single type of chlorenchyma cell. These data indicate that the lamina joint and pulvinus have unique expression patterns of C3 and C4 enzymes, unlike those in C4-type anatomy.