Molecular Genetic Analysis of Pollination-Induced Senescence in Phalaenopsis Orchids

Abstract

The project investigated the molecular genetic and biochemical basis of pollination-induced senescence of Phalaenopsis flowers. This experimental system offered unique advantages in that senescence is strictly regulated by pollination, providing the basis to experimentally initiate and synchronize senescence in populations of flowers. The postpollination syndrome in the Phalaenopsis orchid system was dissected by investigating the temporal and spatial regulation of ACC synthase gene expression. In the stigma, pollen-borne auxin induces the expression of the auxin-regulated ACC synthase (PS-ACS2) gene, resulting in ACC synthesis within 1 h following pollination. Newly formed ACC is oxidized by basal constitutive ACC oxidase to ethylene, which then induces the expression of the ethylene-regulated ACC synthase(PS-ACS1) and oxidase (ACO1) genes for further autocatalytic production of ethylene. It is speculated that during the 6-h period following pollination, emasculation leads to the production or release of a sensitivity factor that sensitizes the cells of the stigma to ethylene. ACC and ethylene molecules are translocated from the stigma to the labellum and perianth where ethylene induces the expression of PS-ACS1 and ACO1 resulting in an increased production of ACC and ethylene. Organ-localized ethylene is responsible for inrolling and senescence of the labellum and perianth. The regulation of ethylene sensitivity and signal transduction events in pollinated flowers was also investigated. The increase in ethylene sensitivity appeared in both the flower column and the perianth, and was detected as early as 4 h after pollination. The increase in ethylene sensitivity following pollination was not dependent on endogenous ethylene production. Application of linoleic and linoleic acids to Phalaenopsis and Dendrobium flowers enhanced their senescence and promoted ethylene production. Several major lipoxygenase pathway products including JA-ME, traumatic acid, trans-2-hexenal and cis-3-hexenol, also enhanced flower senescence. However, lipoxygenase appears to not be directly involved in the endogenous regulation of pollination-induced Phalaenopsis and Dendrobium flower senescence. The data suggest that short-chain saturated fatty acids may be the ethylene "sensitivity factors" produced following pollination, and that their mode of action involves a decrease in the order of specific regions i the membrane lipid bilayer, consequently altering ethylene action. Examination of potential signal transduction intermediates indicate a direct involvement of GTP-binding proteins, calcium ions and protein phosphorylation in the cellular signal transduction response to ethylene following pollination. Modulations of cytosolic calcium levels allowed us to modify the flowers responsiveness to ethylene.