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RNA-Seq Analysis of Laser-Capture Microdissected Cells of the Developing Central Starchy Endosperm of Maize

ABSTRACT: Endosperm is a product of double fertilization, and provides nutrients and signals to the embryo during seed development in flowering plants. Early stages of endosperm development are critical for the development of its storage capacity through synthesis and accumulation of starch and storage proteins. Here we report on the isolation and sequencing of mRNAs from the central portion of the starchy endosperm of Zea mays (maize) B73 at 6 days after pollination. We detected high correlation among the four biological replicates of RNAs isolated using laser-capture microdissection of the cell type. Because the assayed stage of development precedes the synthesis and accumulation of the major storage proteins and starch in the endosperm, our dataset likely include mRNAs for genes that are involved in control and establishment of these developmental programs.

ORGANISM(S): Zea mays

PROVIDER: GSE58504 | GEO | 2014/06/17

SECONDARY ACCESSION(S): PRJNA252770

REPOSITORIES: GEO

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Project description:Barley (Hordeum vulgare) is one of the major food sources for humans and forage source for animal livestock. Barley endosperm is structured into three distinct cell layers: the starchy endosperm, which acts essentially as storage tissue for starch, the subaleurone, which is characterized by a high accumulation of endoplasmic reticulum (ER)-derived seed storage proteins (SSP) and finally the aleurone beside the seed coat with a prominent role during seed germination. Prolamins account for more than 50% of the total protein amount in mature seeds. Together with other seed storage proteins (SSPs) they are important for both grain quality and flour quality. Prolamins are synthesized on the rough ER, translocated into the ER lumen and accumulate in distinct, ER-derived protein bodies (PBs) that are most abundant in the SE. PB formation is regulated by the protein disulfide isomerase (PDI) that is involved in the disulfide transfer pathway. Here, we used laser microdisection (LMD) to characterize spatio-temporal molecular and morphological differences of the ER during barley endosperm development. We revealed by nanoLC-MS/MS proteomic analyses performed on whole seeds and collected tissues at different seed development stages that the protein level of the protein disulfide isomerase HvPDIL1-1 is spatio-temporally regulated in developing barley endosperm. Our microscopic studies showed that HvPDIL1-1 preferentially accumulates in SE, especially at 12 days after pollination (dap). HvPDIL1-1 re-localized from PBs to the protein matrix at the periphery of starch granules along grain filling process. Detailed analysis of SE proteome dynamics identified clusters of proteins with similar expression pattern as HvPDIL1-1, which were analysed in a protein-protein network. It revealed a strong functional interconnection between transcription and translation, protein folding and amino acid synthesis with sucrose and starch metabolism. Our data indicate a role of HvPDIL1-1 in the coordination of protein synthesis and prolamins deposition during grain filling processes in developing barley endosperm. These results are discussed in relation to the putative role of HvPDIL1-1 for cereal food end-product quality and recombinant protein production in cereal seeds.

Project description:This reports the transcript profiling of the aleurone and starchy endosperm layers of wheat seed over 3 time points critical in the development of the aleurone layer. Wheat is a critical food source globally. The aleurone layer develops from the starchy endosperm and is a concentrated source of vitamins and minerals, essential for the germination of the plant embryo. However the molecular mechanisms behind the development of this layer remain poorly understood. Here we present the first direct systematic comparison of the transcriptomes of the aleurone and starchy endosperm tissues of the wheat seed (Triticum aestivum) at time points critical to the development of the aleurone layer of 6, 9 and 14 days post anthesis. Gene expression patterns reflect the changing role of these tissues in seed development. Illumina sequencing gave 25 to 55 million sequence reads per tissue, of the trimmed reads, 70 – 81% mapped to reference expressed sequence transcripts. To quantify transcript abundance, RNA-Seq normalisation was performed to generate RPKM values, these were used in comparative analyses between the tissues at each time point using Kals Z-test. Sequences with significantly different RPKM values were categorised on the basis of tissue and time point expression and functionally annotated using standardised gene ontology vocabularies, revealing two very distinct tissues. In conclusion we show the relationships between and the fundamental biological reprogramming of the two major biologically and economically significant tissues of the wheat seed over time. Understanding these changes in gene expression profiles is essential to mining the potential these tissues hold for human nutrition and contributing to foundational and systems biology of this important crop. Examines the transcript profile of aleurone and starchy endoperm tissues at 6,9 and 14DPA. A minimum of 5 individual seeds from 3 separate spikes from three individual plants were pooled for each tissue preparation.

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RNA-Seq Analysis of Laser-Capture Microdissected Cells of the Developing Central Starchy Endosperm of Maize

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