Tadashi Sakata

Retinoblastoma and Its Binding Partner MSI1 Control Imprinting in Arabidopsis

Parental genomic imprinting causes preferential expression of one of the two parental alleles. In mammals, differential sex-dependent deposition of silencing DNA methylation marks during gametogenesis initiates a new cycle of imprinting. Parental genomic imprinting has been detected in plants and relies on DNA methylation by the methyltransferase MET1. However, in contrast to mammals, plant imprints are created by differential removal of silencing marks during gametogenesis. In Arabidopsis, DNA demethylation is mediated by the DNA glycosylase DEMETER (DME) causing activation of imprinted genes at the end of female gametogenesis. On the basis of genetic interactions, we show that in addition to DME, the plant homologs of the human Retinoblastoma (Rb) and its binding partner RbAp48 are required for the activation of the imprinted genes FIS2 and FWA. This Rb-dependent activation is mediated by direct transcriptional repression of MET1 during female gametogenesis. We have thus identified a new mechanism required for imprinting establishment, outlining a new role for the Retinoblastoma pathway, which may be conserved in mammals.

Drosophila Nedd4 Regulates Endocytosis of Notch and Suppresses Its Ligand-Independent Activation

BACKGROUNDnLigand-induced proteolytic cleavage and internalization of the plasma membrane receptor Notch leads to its activation. Ligand-independent, steady-state internalization of Notch, however, does not lead to activation. The mechanism by which downstream effectors discriminate between these bipartite modes of Notch internalization is not understood. Nedd4 is a HECT domain-containing E3 ubiquitin ligase that targets transmembrane receptors containing the PPSY motif for endocytosis. Deltex is a positive Notch signaling regulator that encodes a putative ubiquitin ligase of the ring finger type.nnnRESULTSnWe used the Drosophila system to show that Notch is ubiquitinated and destabilized by Nedd4 in a manner requiring the PPSY motif in the Notch intracellular domain. Loss of Nedd4 function dominantly suppresses the Notch and Deltex mutant phenotypes, and its hyperactivation attenuates Notch activity. In tissue culture cells, the dominant-negative form of Nedd4 blocks steady-state Notch internalization and activates Notch signaling independently of ligand binding. This effect was further potentiated by Deltex. Nedd4 destines Deltex for degradation in a Notch-dependent manner.nnnCONCLUSIONSnNedd4 antagonizes Notch signaling by promoting degradation of Notch and Deltex. This Nedd4 function may be important for protecting unstimulated cells from sporadic activation of Notch signaling.

Auxins reverse plant male sterility caused by high temperatures

Auxin levels are well regulated in cells and tissues by both transport and local biosynthesis, and its distribution is important for the modulation of cell proliferation, differentiation, development, tropisms and high-temperature response. Activation of auxin biosynthesis with increased temperatures reported in certain plant tissues. In contrast, our studies indicated that male tissue-specific auxin reduction via transcriptional repression of the YUCCA auxin biosynthesis genes is the primary cause of high temperature injury, which leads the abortion of pollen development in Arabidopsis and barley Hordeum vulgare L. Furthermore, the abortion can be reversed by the application of exogenous auxin, suggesting that the application may maintain crop yields during the current global warming crisis.With global warming, plant high temperature injury is becoming an increasingly serious problem. In wheat, barley, and various other commercially important crops, the early phase of anther development is especially susceptible to high temperatures. Activation of auxin biosynthesis with increased temperatures has been reported in certain plant tissues. In contrast, we here found that under high temperature conditions, endogenous auxin levels specifically decreased in the developing anthers of barley and Arabidopsis. In addition, expression of the YUCCA auxin biosynthesis genes was repressed by increasing temperatures. Application of auxin completely reversed male sterility in both plant species. These findings suggest that tissue-specific auxin reduction is the primary cause of high temperature injury, which leads to the abortion of pollen development. Thus, the application of auxin may help sustain steady yields of crops despite future climate change.

Effects of High Temperature on the Development of Pollen Mother Cells and Microspores in Barley Hordeum vulgare L.

The development of the inflorescence, microspores and anthesis were well synchronized among individuals or in the panicles of barley under controlled environmental conditions. To study the effects of high-temperature stress on the development of pollen mother cells (PMCs) and microspores, the plants were subjected to high temperature treatment at different stages of reproductive growth. When plants were exposed to high temperature for five days at the early differentiation stage of the panicle, pollen grains had apparently normal exine but no or little cytoplasm. At the pre-meiotic stage of PMCs, high temperature caused subsequent development of short anthers possessing no pollen grains. When plants were exposed to high temperature during meiosis of PMCs, all pollen grains possessed exine and were swollen but showed little starch accumulation. In these plants treated at high temperature, the panicles at the heading stage had a normal appearance, but their seeds were virtually sterile. These results indicated that there are at least three stages of reproductive growth hypersensitive to high temperature, which resulted in abnormal terminal phenotypes different from one another.

High-temperature induction of male sterility during barley (Hordeum vulgare L.) anther development is mediated by transcriptional inhibition

High-temperature induction of male sterility during floral organogenesis is a critical problem for barley (Hordeum vulgare L.) crops that the molecular basis is incompletely understood. Gene expression and differentiation of anthers were examined under normal (20°C day/15°C night) and elevated (30°C day/25°C night) temperatures. Serial analysis of gene expression analysis displayed contrasting profiles of gene expression in early panicles between control and high-temperature conditions. Several transcripts dramatically upregulated before development and differentiation of anther wall layers in normal temperatures, including histone H3, H4 and glycine-rich RNA-binding protein genes, were not upregulated at elevated temperatures and typically abundant mRNAs, such as 60S ribosomal protein L27a and glyoxalase I, appeared to be downregulated. Instead, development and differentiation of tapetum cells and pollen mother cells were completely aborted. Failure of transcriptional reactivation with return to normal temperatures increases with duration of elevated temperatures and is strongly correlated with observation of male sterility. Hyper-phosphorylation of the ser-5 residue of the C-terminal domain of the largest subunit of RNA polymerase II (RPB1) was noted to occur under high-temperature conditions. These results indicate that early development and differentiation of barley anthers are very sensitive to high-temperature stress causing major alterations in gene expression.

The two male gametes share equal ability to fertilize the egg cell in Arabidopsis thaliana.

Summary The seed of a flowering plant develops from an ovule containing two distinct female gametes — the egg cell and the central cell — that are fertilized by a pair of non-motile sperm cells conveyed by the pollen tube. With a few exceptions [1], the two sperm cells, derived from a symmetrical mitosis, are isomorphic and seem to express a similar gene repertoire [2]. Since the discovery of double fertilization in flowering plants at the end of the 19th century, it has been a long standing question whether the two sperm cells are functionally equivalent, that is, whether they are capable of fertilizing the egg cell and the central cell in equal measure.

Reduction of Gibberellin by Low Temperature Disrupts Pollen Development in Rice

Gibberellins and expression levels of their biosynthesis genes decrease in developing anthers on exposure to moderate low temperatures, disrupting pollen development and reducing grain yields. Microsporogenesis in rice (Oryza sativa) plants is susceptible to moderate low temperature (LT; approximately 19°C) that disrupts pollen development and causes severe reductions in grain yields. Although considerable research has been invested in the study of cool-temperature injury, a full understanding of the molecular mechanism has not been achieved. Here, we show that endogenous levels of the bioactive gibberellins (GAs) GA4 and GA7, and expression levels of the GA biosynthesis genes GA20ox3 and GA3ox1, decrease in the developing anthers by exposure to LT. By contrast, the levels of precursor GA12 were higher in response to LT. In addition, the expression of the dehydration-responsive element-binding protein DREB2B and SLENDER RICE1 (SLR1)/DELLA was up-regulated in response to LT. Mutants involved in GA biosynthetic and response pathways were hypersensitive to LT stress, including the semidwarf mutants sd1 and d35, the gain-of-function mutant slr1-d, and gibberellin insensitive dwarf1. The reduction in the number of sporogenous cells and the abnormal enlargement of tapetal cells occurred most severely in the GA-insensitive mutant. Application of exogenous GA significantly reversed the male sterility caused by LT, and simultaneous application of exogenous GA with sucrose substantially improved the extent of normal pollen development. Modern rice varieties carrying the sd1 mutation are widely cultivated, and the sd1 mutation is considered one of the greatest achievements of the Green Revolution. The protective strategy achieved by our work may help sustain steady yields of rice under global climate change.

Tissue-specific auxin signaling in response to temperature fluctuation

Auxin levels are well regulated in cells and tissues by both transport and local biosynthesis, and its distribution is important for the modulation of cell proliferation, differentiation, development, tropisms and high-temperature response. Activation of auxin biosynthesis with increased temperatures reported in certain plant tissues. In contrast, our studies indicated that male tissue-specific auxin reduction via transcriptional repression of the YUCCA auxin biosynthesis genes is the primary cause of high temperature injury, which leads the abortion of pollen development in Arabidopsis and barley Hordeum vulgare L. Furthermore, the abortion can be reversed by the application of exogenous auxin, suggesting that the application may maintain crop yields during the current global warming crisis.