Ethylene receptors are multispanning membrane proteins that negatively regulate ethylene responses via the formation of a signaling complex with downstream elements. To better understand their biochemical functions, we investigated the membrane topology and subcellular localization of CmERS1, a melon (Cucumis melo) ethylene receptor that has three putative transmembrane domains at the N terminus. Analyses using membrane fractionation and green fluorescent protein imaging approaches indicate that CmERS1 is predominantly associated with the endoplasmic reticulum (ER) membrane. Detergent treatments of melon microsomes showed that the receptor protein is integrally bound to the ER membrane. A protease protection assay and N-glycosylation analysis were used to determine membrane topology. The results indicate that CmERS1 spans the membrane three times, with its N terminus facing the luminal space and the large C-terminal portion lying on the cytosolic side of the ER membrane. This orientation provides a platform for interaction with the cytosolic signaling elements. The three N-terminal transmembrane segments were found to function as topogenic sequences to determine the final topology. High conservation of these topogenic sequences in all ethylene receptor homologs identified thus far suggests that these proteins may share the same membrane topology.
Summary• Ethylene evolution from plants inhibits Agrobacterium-mediated genetic transformation, but the mechanism is little understood. In this study, the possible role of ethylene in Agrobacterium-mediated genetic transformation was clarified.• It was tested whether or not plant ethylene sensitivity affected genetic transformation; the sensitivity might regulate bacterial growth during co-cultivation and vir gene expression in Agrobacterium tumefaciens. For these experiments, melon (Cucumis melo) was used, in which ethylene sensitivity was controlled by chemicals, and Arabidopsis ethylene-insensitive mutants.• Agrobacterium-mediated genetic transformation was inhibited in ethylene-sensing melon, whereas, in Arabidopsis ethylene-insensitive mutant, it was enhanced. However, the ethylene sensitivity did not affect bacterial growth. vir gene expression was inhibited by application of plant exudate from ethylene-sensitive plants. The inhibitory effect of the ethylene sensitivity on genetic transformation relieved the activation of vir gene expression in A. tumefaciens with vir gene inducer molecule (acetosyringone, AS) or A. tumefaciens mutant strain which has constitutive vir gene expression.• These results indicate that ethylene evolution from a plant inoculated with A. tumefaciens inhibited vir gene expression in A. tumefaciens through the ethylene signal transduction in the plant, and, as a result, Agrobacterium-mediated genetic transformation was inhibited.
Four sand units deposited by tsunamis and one sand unit deposited by storm surge(s) were identified in a muddy marsh succession in a narrow coastal lowland along the Pacific coast of central Japan. Tsunamis in ad 1498, 1605, 1707 and 1854 that were related to large subduction‐zone earthquakes along the Nankai Trough, and storm surges in 1680 and/or 1699 were responsible for the deposition of these sand units. These sand units are distinguished by lithofacies, sedimentary structures, grain‐size and mineral composition, and radiocarbon ages; their ages are supported by events in local historical records. The tsunami deposits in the study area are massive or parallel‐laminated sands, with associated intraclasts, gravels, draping mud layers and, rarely, a return‐flow subunit. The storm surge deposits are devoid of these characteristics, and are composed of groups of thin, current ripple‐laminated sand layers. The differences in sedimentary structures between the tsunami and storm surge deposits are attributed to the different characteristics of tsunami and storm waves.
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