Identification of signalling components and regulatory pathways determining development of the barley endosperm

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In barley grains the coordinated development of endosperm transfer cells (ETCs) is responsible for the regular differentiation of the endosperm resulting in two major cell types: aleurone and starchy endosperm, which includes the prismatic endosperm (facing ETCs) and the starchy endosperm of the wing regions. The barley endosperm mutant seg8, which is defective in cellularisation and differentiation of ETCs, reveals reduced or missing prismatic endosperm and 70% reduction of seed weight. Thus, seg8 is a suitable model for the identification of components determining endosperm development. One objective of the proposal is to identify the mutated seg8 gene by using genomic barley sequence information. In addition, comparative transcript, metabolite and hormone profiling will be performed on microdissected ETCs and prismatic endosperm cells of seg8 and wild type (WT) grains. There is strong evidence that changes in seg8 grains are related to altered ABA functions. Thus, another objective is to study ABA biosynthesis, signalling and distribution as related to endosperm cellularisation and differentiation in seg8 and WT grains. Expression of ABA biosynthesis genes and signalling components will be studied by mRNA in situ hybridisation and qRT-PCR. Those regions showing altered patterns of ABA biosynthesis in seg8 will be analysed by transcript and metabolite profiling after microdissection in comparison to the WT. Bioinformatics network analysis will be used to identify signalling cascades regulating endosperm development.