Barley endosperm: stress response of endomembranes and ESCRT

Drought, heat, high salt and low temperature are the most common environmental stress factors that influence plant growth and development, and are major limits on plant productivity in cultivated areas worldwide. Abiotic stress thus has a strong impact on crop productivity and on central developmental functions of the caryopsis. Heat, drought, and salt induce different subcellular effects. Besides altered biochemical pathways, the endomembrane system is affected by abiotic stress. Above all, vacuolar compartmentalization and vesicle trafficking have been reported. The Endosomal sorting complex required for transport (ESCRT) consists of four subunits 0, I, II, and III and is responsible for endocytic recycling of plasma membrane proteins. ESCRT proteins are strongly conserved and are involved in many different cellular processes. It was shown that ESCRT proteins are indispensable for maize endosperm development and in ESCRT-mediated endosomal trafficking for salt- and drought tolerance. Recently we have identified barley ESCRT-III members in the (model) crop Hordeum vulgare (Hv) and show that all known members are expressed in developing barley endosperm. Our initial studies of ESCRT-III in barley endosperm showed that the localization and involvement of an ESCRT-III protein in diverse trafficking pathways is depending on the cell layer. This result is accompanied by the spatio-temporal endomembrane rearrangement in developing barley endosperm, especially between 8 and 12 days after pollination. This is the first study to undertake a wide-ranging analysis of the barley endomembrane system in response to drought, heat and salt stimuli. Endocytosis regulating-, ESCRT-III and ESCRT-III interacting proteins will be analysed in response to drought, heat and salt stimuli during barley endosperm development and germination. In detail, the localization of fluorescently tagged proteins in the endosperm of transgenic lines will be studied in response to stress. Furthermore, the endomembrane system of knock-down lines will be analysed in response to stress. Stress stimuli will be performed starting at 8 dap of endosperm development and during germination in plant incubators and caryopses will be analysed by confocal microscopy. A particular strength of this work will be the spatio-temporal resolution achieved by laser dissection microscopy. The multidisciplinary study using cell layer-specific subcellular live cell imaging, transcriptomics and proteomics is a strong approach to determine the interplay of endocytosis and ESCRT-III in response to abiotic stress in barley endosperm and will contribute to the understanding of mechanisms important to genetic improvement of crop plants.