Comprehensive Network Analysis of Anther-Expressed Genes in Rice by the Combination of 33 Laser Microdissection and 143 Spatiotemporal Microarrays

Bioscience and Biotechnology Center, Nagoya University, Nagoya, Japan.
PLoS ONE (Impact Factor: 3.53). 10/2011; 6(10):e26162. DOI: 10.1371/journal.pone.0026162
Source: PubMed

ABSTRACT Co-expression networks systematically constructed from large-scale transcriptome data reflect the interactions and functions of genes with similar expression patterns and are a powerful tool for the comprehensive understanding of biological events and mining of novel genes. In Arabidopsis (a model dicot plant), high-resolution co-expression networks have been constructed from very large microarray datasets and these are publicly available as online information resources. However, the available transcriptome data of rice (a model monocot plant) have been limited so far, making it difficult for rice researchers to achieve reliable co-expression analysis. In this study, we performed co-expression network analysis by using combined 44 K agilent microarray datasets of rice, which consisted of 33 laser microdissection (LM)-microarray datasets of anthers, and 143 spatiotemporal transcriptome datasets deposited in RicexPro. The entire data of the rice co-expression network, which was generated from the 176 microarray datasets by the Pearson correlation coefficient (PCC) method with the mutual rank (MR)-based cut-off, contained 24,258 genes and 60,441 genes pairs. Using these datasets, we constructed high-resolution co-expression subnetworks of two specific biological events in the anther, "meiosis" and "pollen wall synthesis". The meiosis network contained many known or putative meiotic genes, including genes related to meiosis initiation and recombination. In the pollen wall synthesis network, several candidate genes involved in the sporopollenin biosynthesis pathway were efficiently identified. Hence, these two subnetworks are important demonstrations of the efficiency of co-expression network analysis in rice. Our co-expression analysis included the separated transcriptomes of pollen and tapetum cells in the anther, which are able to provide precise information on transcriptional regulation during male gametophyte development in rice. The co-expression network data presented here is a useful resource for rice researchers to elucidate important and complex biological events.

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    ABSTRACT: Plants lack a germ line; consequently, during reproduction adult somatic cells within flowers must switch from mitotic proliferation to meiosis. In maize (Zea mays L.) anthers, hypoxic conditions in the developing tassel trigger pre-meiotic competence in the column of pluripotent progenitor cells in the center of anther lobes, and within 24 hr these newly specified germinal cells have patterned their surrounding neighbors to differentiate as the first somatic niche cells. Transcriptomes were analyzed by microarray hybridization in carefully staged whole anthers during initial specification events, after the separation of germinal and somatic lineages, during the subsequent rapid mitotic proliferation phase, and during final pre-meiotic germinal and somatic cell differentiation. Maize anthers exhibit a highly complex transcriptome constituting nearly three-quarters of annotated maize genes, and expression patterns are dynamic. Laser microdissection was applied to begin assigning transcripts to tissue and cell types and for comparison to transcriptomes of mutants defective in cell fate specification. Whole anther proteomes were analyzed at three developmental stages by mass spectrometric peptide sequencing using size-fractionated proteins to evaluate the timing of protein accumulation relative to transcript abundance. New insights include early and sustained expression of meiosis-associated genes (77.5% of well-annotated meiosis genes are constitutively active in 0.15 mm anthers), an extremely large change in transcript abundances and types a few days before meiosis (including a class of 1340 transcripts absent specifically at 0.4 mm), and the relative disparity between transcript abundance and protein abundance at any one developmental stage (based on 1303 protein-to-transcript comparisons).
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    ABSTRACT: Anther-specific chalcone synthase-like enzyme (ASCL), an early evolved plant type III polyketide synthase (PKS), is involved in the biosynthesis of sporopollenin, the stable biopolymer found in the exine layer of the wall of a spore or pollen grain. HpPKS1 from Hypericum perforatum was previously shown to be expressed mainly in young flower buds but also in leaf and other tissues at lower levels. Angiosperm ASCLs, identified by sequence and phylogenetic analyses, are divided into two sister clades, the Ala-clade and Val-clade, and HpPKS1 belongs to the Ala-clade. Recombinant HpPKS1 produced triketide and, to a lesser extent, tetraketide alkylpyrones from medium (C6)- to very long-chain (C24) fatty acyl-CoA substrates. Similar to other ASCLs, HpPKS1 also preferred hydroxyl fatty acyl-CoA esters over the analogous unsubstituted fatty acyl-CoA esters. To study the structural basis of the substrate preference, mutants of Ala200 and Ala215 at the putative active site and Arg202 and Asp211 at the modeled acyl-binding tunnel were constructed. The A200T/A215Q mutant accepted decanoyl-CoA, a poor substrate for the wild-type enzyme, possibly due to active site constriction by bulkier substitutions. The substrate preference of the A215V and A200T/A215Q mutants shifted toward non-hydroxylated, medium- to long-chain fatty acyl-CoA substrates. The R202L/D211V double mutant was selective for acyl-CoA with chain lengths of C16–C18 and showed diminished preference for the hydroxylated acyl-CoA substrates. Transient up-regulation by abscisic acid and down-regulation by jasmonic acid and wounding suggested that HpPKS1 and possibly other Ala-clade ASCLs may be involved in the biosynthesis of minor cell wall components in non-anther tissues.
    FEBS Journal 07/2014; 281(17). DOI:10.1111/febs.12920 · 3.99 Impact Factor
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