Unknown,Transcriptomics,Genomics,Proteomics

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Dynamic transcript abundance during cellularisation of the developing barley endosperm: Temporal regulation of cell wall synthesis

ABSTRACT: Grain development in the Poaceae defines important end-use properties such as yield, quality and nutritive value. Microarray analyses have been performed on barley grain endosperm extracts from three to eight days after pollination (DAP), when cellularization of the syncytium occurs through the growth of cell walls around individual nuclei. Profiling of transcripts differentially expressed over time reveal 56 specific modules of genes that cluster into 15 groups. Expression patterns have been superimposed upon microscopy data, which identify the timing of key stages in grain development. Thus, cellularization is complete at six DAP, aleurone-related genes can be detected at seven to eight DAP, and starch synthase and hordein genes increase dramatically at seven and eight DAP, respectively. Genes known to be involved in cell wall metabolism are found predominantly in a single module, but analysis using a gene ontology approach splits these genes into four modules, which remain in a single cluster. Transcript levels of the cell wall-related genes peak at seven DAP and the developmental patterns of genes involved in arabinoxylan and (1,3;1,4)-β-glucan synthesis are defined. The transcript data are publicly available (www.etc.) and can be used to interrogate co-expression and differential expression patterns for other groups of genes. In addition, the examination of transcription factor genes that are co-expressed in modules of genes involved in specific processes, such as aleurone differentiation, can be used to identify candidate genes for the control of those particular processes during barley grain development.

ORGANISM(S): Hordeum vulgare

SUBMITTER: Pete Hedley

PROVIDER: E-MTAB-2728 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Project description:Hordeum vulgare (barley) hordoindolines (HINs), HINa, HINb1 and HINb2, are orthologous proteins of wheat puroindolines (PINs) that are small, basic, cysteine-rich seed-specific proteins and responsible for grain hardness. Grain hardness, is, next to its protein content, a major quality trait. In barley, HINb is most highly expressed in the mid-stage developed endosperm and is associated with both major endosperm texture and grain hardness. However, data required tounderstand the spatio-temporal dynamics of HIN transcripts and HIN protein regulation during grain filling processes are missing. Using reverse transcription quantitative PCR (RT-qPCR) and proteomics we analyzed HIN transcript and HIN protein abundance from whole seeds (WSs) at four ((6 days after pollination (dap), 10 dap, 12 dap and ≥ 20 dap)) as well as from aleurone, subaleurone and starchy endosperm at two (12 dap and ≥ 20 dap) developmental stages. At the WS level, results from RT-qPCR, proteomics and western blot showed a continuous increase of HIN transcript and HIN protein abundance across these four developmental stages. Miroscopic studies revealed HIN localization mainly at the vacuolar membrane in the aleurone, at protein bodies (PBs) in subaleurone and at the periphery of starch granules in the starchy endosperm. Laser microdissetion (LMD) proteomic analyses identified HINb2 as the most prominent HIN protein in starchy endosperm at ≥ 20 dap. Additionally, our quantification data revealed a poor correlation between transcript and protein levels of HINs in subaleurone during development. Here, we correlated data achieved by RT-qPCR, proteomics and microscopy that reveal different expression and localization pattern of HINs in each layer during barley endosperm development. This indicats a contribution of each tissue to the regulation of HINs during grain filling. The effect of the high protein abundance of HINs in the starchy endosperm and their localization at the periphery of starch granules at late development stages at the high end-product quality is discussed. Understanding the spatio-temporal regulated HINs is essential to improve barley quality traits for high end-product quality, as hard texture of the barley grain is regulated by the ratio between HINb/HINa.