Differential responses of anthers of stress tolerant and sensitive wheat cultivars to high temperature stress.

MAIN CONCLUSION: Transcriptomic analyses identified anther-expressed genes in wheat likely to contribute to heat tolerance and hence provide useful genetic markers. The genes included those involved in hormone biosynthesis, signal transduction, the heat shock response and anther development. Pollen development is particularly sensitive to high temperature heat stress. In wheat, heat-tolerant and heat-sensitive cultivars have been identified, although the underlying genetic causes for these differences are largely unknown. The effects of heat stress on the developing anthers of two heat-tolerant and two heat-sensitive wheat cultivars were examined in this study. Heat stress (35 °C) was found to disrupt pollen development in the two heat-sensitive wheat cultivars but had no visible effect on pollen or anther development in the two heat-tolerant cultivars. The sensitive anthers exhibited a range of developmental abnormalities including an increase in unfilled and clumped pollen grains, abnormal pollen walls and a decrease in pollen viability. This subsequently led to a greater reduction in grain yield in the sensitive cultivars following heat stress. Transcriptomic analyses of heat-stressed developing wheat anthers of the four cultivars identified a number of key genes which may contribute to heat stress tolerance during pollen development. Orthologs of some of these genes in Arabidopsis and rice are involved in regulation of the heat stress response and the synthesis of auxin, ethylene and gibberellin. These genes constitute candidate molecular markers for the breeding of heat-tolerant wheat lines.

The MYB80 transcription factor is required for pollen development and the regulation of tapetal programmed cell death in Arabidopsis thaliana.

Arabidopsis thaliana MYB80 (formerly MYB103) is expressed in the tapetum and microspores between anther developmental stages 6 and 10. MYB80 encodes a MYB transcription factor that is essential for tapetal and pollen development. Using microarray analysis of anther mRNA, we identified 404 genes differentially expressed in the myb80 mutant. Employing the glucocorticoid receptor system, the expression of 79 genes was changed when MYB80 function was restored in the myb80 mutant following induction by dexamethasone. Thirty-two genes were analyzed using chromatin immunoprecipitation, and three were identified as direct targets of MYB80. The genes encode a glyoxal oxidase (GLOX1), a pectin methylesterase (VANGUARD1), and an A1 aspartic protease (UNDEAD). All three genes are expressed in the tapetum and microspores. Electrophoretic mobility shift assays confirmed that MYB80 binds to all three target promoters, with the preferential binding site containing the CCAACC motif. TUNEL assays showed that when UNDEAD expression was silenced using small interfering RNA, premature tapetal and pollen programmed cell death occurred, resembling the myb80 mutant phenotype. UNDEAD possesses a mitochondrial targeting signal and may hydrolyze an apoptosis-inducing protein(s) in mitochondria. The timing of tapetal programmed cell death is critical for pollen development, and the MYB80/UNDEAD system may regulate that timing.

MYB80, a regulator of tapetal and pollen development, is functionally conserved in crops.

The Arabidopsis AtMYB80 transcription factor (formerly AtMYB103) regulate genes essential for tapetal and pollen development. One of these genes, coding for an aspartic protease (UNDEAD), may control the timing of tapetal programmed cell death (PCD). In crop plants such as rice and wheat, abiotic stresses lead to abnormal tapetal development resulting in delayed PCD. Manipulation of AtMYB80 function has been used to develop a reversible male sterility system applicable to hybrid crop production. MYB80 homologs were cloned from wheat, rice, canola and cotton. The promoters of the homologs drove temporal and spatial expression patterns of the GUS reporter gene in the tapetum and microspores of Arabidopsis anthers identical to the AtMYB80 promoter. A short region is conserved in all five MYB80 promoters. The MYB80 homolog genes, driven by the AtMYB80 or their respective promoters, rescued the atmyb80 mutant, completely restoring male fertility. The canola MYB80 was fused to the EAR (ERF-associated amphiphilic repression) repressor and canola plants transgenic for the construct exhibited premature tapetal degradation and subsequent pollen abortion. The five MYB80 homologs all shared a 44 amino acid sequence immediately adjacent to the R2R3 domain which appears to be necessary for MYB80 function.

Anther Morphological Development and Stage Determination in Triticum aestivum.

Anther development progresses through 15 distinct developmental stages in wheat, and accurate determination of anther developmental stages is essential in anther and pollen studies. A detailed outline of the development of the wheat anther through its entire developmental program, including the 15 distinct morphological stages, is presented. In bread wheat (Triticum aestivum), anther developmental stages were correlated with five measurements, namely auricle distance, spike length, spikelet length, anther length and anther width. Spike length and auricle distance were shown to be suitable for rapid anther staging within cultivars. Anther length is an accurate measurement in determining anther stages and may be applicable for use between cultivars. Tapetal Programmed Cell Death (PCD) in wheat begins between late tetrad stage (stage 8) and the early young microspore stage (stage 9) of anther development. Tapetal PCD continues until the vacuolate pollen stage (stage 11), at which point the tapetum fully degrades. The timing of tapetal PCD initiation is slightly delayed compared to that in rice, but is two stages earlier than in the model dicot Arabidopsis. The MYB80 gene, which encodes a transcription factor regulating the timing of tapetal PCD, reaches its peak expression at the onset of tapetal PCD in wheat.

Layers of regulation - Insights into the role of transcription factors controlling mucilage production in the Arabidopsis seed coat.

A polysaccharide-rich mucilage is released from the seed coat epidermis of numerous plant species and has been intensively studied in the model plant Arabidopsis. This has led to the identification of a large number of genes involved in the synthesis, secretion and modification of cell wall polysaccharides such as pectin, hemicellulose and cellulose being identified. These genes include a small network of transcription factors (TFs) and transcriptional co-regulators, that not only regulate mucilage production, but epidermal cell differentiation and in some cases flavonoid biosynthesis in the internal endothelial layer of the seed coat. Here we focus on the function of these regulators and propose a simplified model where they are assigned to a hierarchical gene network with three regulatory levels (tiers) as a means of assisting in the interpretation of the complexity. We discuss limitations of current methodologies and highlight some of the problems associated with defining the function of TFs, particularly those that perform different functions in adjacent layers of the seed coat. We suggest approaches that should provide a more accurate picture of the function of transcription factors involved with mucilage production and release.

Tapetal development and abiotic stress: a centre of vulnerability.

Many self-fertilising crops are particularly sensitive to abiotic stress at the reproductive stage. In rice (Oryza sativa L.) and wheat (Triticum aestivum L.), for example, abiotic stress during meiosis and the young microspore stage indicates the tapetum is highly vulnerable and that the developmental program appears to be compromised. Tapetal hypertrophy can occur as a consequence of cold and drought stress, and programmed cell death (PCD) is delayed or inhibited. Since the correct timing of tapetal PCD is essential for pollen reproduction, substantial losses in grain yield occur. In wheat and rice, a decrease in tapetal cell wall invertase levels is correlated with pollen abortion and results in the amount of hexose sugars reaching the tapetum, and subsequently the developing microspores, being severely reduced ('starvation hypothesis'). ABA and gibberellin levels may be modified by cold and drought, influencing levels of cell wall invertase(s) and the tapetal developmental program, respectively. Many genes regulating tapetal and microspore development have been identified in Arabidopsis thaliana (L.) Heynh. and rice and the specific effects of abiotic stresses on the program and pathways can now begin to be assessed.