Arabidopsis TSO1 regulates directional processes in cells during floral organogenesis.

Flowers of the previously described Arabidopsis tso1-1 mutant had aberrant, highly reduced organs in place of petals, stamens, and carpels. Cells of tso1-1 flowers had division defects, including failure in cytokinesis, partial cell wall formation, and elevated nuclear DNA content. We describe here two new tso1 alleles (tso1-3 and tso1-4), which caused defects in ovule development, but had little effect on gross floral morphology. Early ovule development occurred normally in tso1-3 and tso1-4, but the shapes and alignments of integument cells became increasingly more disordered as development progressed. tso1-3 ovules usually lacked embryo sacs due to a failure to form megaspore mother cells. The cell division defects described for the strong tso1-1 mutant were rarely observed in tso1-3 ovules. The aberrations in tso1-3 mutants primarily resulted from a failure in directional expansion of cells and/or coordination of this process among adjacent cells. Effects of tso1-3 appeared to be independent of effects of other ovule development mutations, with the exception of leunig, which exhibited a synergistic interaction. The data are consistent with TSO1 acting in processes governing directional movement of cellular components, indicating a likely role for TSO1 in cytoskeletal function.     

Regulation of meristem organization and cell division by TSO1, an Arabidopsis gene with cysteine-rich repeats

In higher plants, meristem organization and cell division regulation are two fundamentally important and intimately related biological processes. Identifying and isolating regulatory genes in these processes is essential for understanding higher plant growth and development. We describe the molecular isolation and analyses of an Arabidopsis gene, TSO1, which regulates both of these processes. We previously showed that tso1 mutants displayed defects in cell division of floral meristem cells including partially formed cell walls, increased DNA content, and multinucleated cells (Liu, Z., Running, M. P. and Meyerowitz, E. M. (1997). Development 124, 665-672). Here, we characterize a second defect of tso1 in influorescence meristem development and show that the enlarged influorescence in tso1 mutants results from repeated division of one inflorescence meristem into two or more influorescence meristems. Using a map-based approach, we isolated the TSO1 gene and found that TSO1 encodes a protein with cysteine-rich repeats bearing similarity to Drosophila Enhancer of zeste and its plant homologs. In situ TSO1 mRNA expression pattern and the nuclear localization of TSO1-GFP are consistent with a regulatory role of TSO1 in floral meristem cell division and in influorescence meristem organization.     

Arabidopsis HAP2 (GCS1) is a sperm-specific gene required for pollen tube guidance and fertilization

In flowering plants, sperm cells develop in the pollen cytoplasm and are transported through floral tissues to an ovule by a pollen tube, a highly polarized cellular extension. After targeting an ovule, the pollen tube bursts, releasing two sperm that fertilize an egg and a central cell. Here, we identified the gene encoding Arabidopsis HAP2, demonstrating that it is allelic to GCS1. HAP2 is expressed only in the haploid sperm and is required for efficient pollen tube guidance to ovules. We identified an insertion (hap2-1) that disrupts the C-terminal portion of the protein and tags mutant pollen grains with the beta-glucuronidase reporter. By monitoring reporter expression, we showed that hap2-1 does not diminish pollen tube length in vitro or in the pistil, but it reduces ovule targeting by twofold. In addition, we show that the hap2 sperm that are delivered to ovules fail to initiate fertilization. HAP2 is predicted to encode a protein with an N-terminal secretion signal, a single transmembrane domain and a C-terminal histidine-rich domain. These results point to a dual role for HAP2, functioning in both pollen tube guidance and in fertilization. Moreover, our findings suggest that sperm, long considered to be passive cargo, are involved in directing the pollen tube to its target.     

Variation in restorer genes and primary sexual investment in gynodioecious Plantago coronopus: the trade-off between male and female function

In many gynodioecious species the nuclear inheritance of male fertility is complex and involves multiple (restorer) genes. In addition to restoring plants from the female (male sterile) to the hermaphrodite (male fertile) state, these genes are also thought to play a role in the determination of the quantity of pollen produced by hermaphrodites. The more restorer alleles a hermaphroditic plant possesses, the higher the pollen production. To test this hypothesis I combined the results of crossing studies of the genetics of male sterility with phenotypic data on investment in stamens and ovules among the progeny of plants involved in these studies. The sex ratio (i.e. the frequency of hermaphrodites among the progeny), being a measure of the number of restorer alleles of the maternal plant, was positively related to the investment in pollen (male function), but negatively related to the investment in ovules (female function), in both field and greenhouse experiments. Consequently, a negative correlation between male and female function was observed (trade-off) and it is suggested that antagonistic pleiotropic effects of restorer genes might be the cause. Phenotypic gender, a measure combining investment in both pollen and ovules, was highly repeatable between field and greenhouse, indicating genetic determination of a more male- or female-biased allocation pattern among the studied plants.     

The higher plant Arabidopsis thaliana encodes a functional CDC48 homologue which is highly expressed in dividing and expanding cells.

We have identified an Arabidopsis thaliana CDC48 gene which, unlike the putative mammalian homologue vasolin-containing protein (VCP), functionally complements Saccharomyces cerevisiae cdc48 mutants. CDC48 is an essential gene in S. cerevisiae and genetic studies suggest a role in spindle pole body separation. Biochemical studies link VCP function to membrane trafficking and signal transduction. We have described the AtCDC48 expression pattern in a multicellular eukaryote; the zones of cell division, expansion and differentiation are physically separated in higher plants, thus allowing the analysis of in situ expression patterns with respect to the state of cell proliferation. AtCDC48 is highly expressed in the proliferating cells of the vegetative shoot, root, floral inflorescence and flowers, and in rapidly growing cells. AtCDC48 mRNA and the encoded protein are up-regulated in the developing microspores and ovules. AtCDC48 expression is down-regulated in most differentiated cell types. AtCDC48p was primarily localized to the nucleus and, during cytokinesis, to the phragmoplast, a site where membrane vesicles are targeted in the deposition of new cell wall materials. This study shows that the essential cell division function of CDC48 has been conserved by, at least, some multicellular eukaryotes and suggests that in higher plants, CDC48 functions in cell division and growth processes.     

Epigenetic silencing of Notch signaling in gastrointestinal cancers

The Notch signaling pathway drives proliferation, differentiation, apoptosis, cell fate choices and maintenance of stem cells during embryogenesis and in self-renewing tissues of the adult. In addition, aberrant Notch signaling has been implicated in several tumors, where Notch can function both as an oncogene or a tumor-suppressor gene, depending on the context. This Extra View aims to review what is currently known about Notch signaling, in particular in gastrointestinal tumors, providing a summary of our data on Notch1 signaling in gastric cancer with results obtained in colorectal cancer (CRC). We have already reported that the epigenetic regulation of the Notch ligand DLL1 controls Notch1 signaling activation in gastric cancer, and that Notch1 inhibition is associated with the diffuse type of gastric cancer. Here, we describe additional data showing that in CRC cell lines, unlike gastric cancer, DLL1 expression is not regulated by promoter methylation. Moreover, in CRC, Notch1 receptor is not affected by any mutation. These data suggest a different regulation of Notch1 signaling between gastric cancer and CRC.     

Characterization of a cytokinesis defective (cyd1) mutant of Arabidopsis.

Although several mutations and genes affecting plant cytokinesis have been identified, mutant screens are not yet saturated and knowledge about gene function is still limited. A novel Arabidopsis mutation, cytokinesis defective1 (cyd1), was identified by partial or missing cell walls in stomata. Stomata with incomplete or no cytokinesis still differentiate and some contain swellings of the outer wall not found in the wild type. The incomplete walls are correctly placed opposite stomatal wall thickenings suggesting that the mutation interferes with the execution of cytokinesis rather than with the placement of the division site. Cytokinesis defects are also detectable in other cell types throughout the plant, defects which include cell wall protrusions, two or more nuclei in one cell, and reduced cell number. The extent of cytokinetic partitioning correlates with nuclear number in abnormal stomata. Many cyd1 epidermal cells, stomata and pollen are larger, and trichomes have more branches. cyd1 is partially lethal with poor seed set and some defective ovules, but many plants are fertile despite abnormalities in vegetative and reproductive development such as missing, reduced, fused or misshapen leaves and floral organs. cyd1 appears to be the only cytokinesis mutant described where defects are known to occur in both mature vegetative and reproductive organs. Thus, the CYD1 gene product appears to be necessary for the execution of cytokinesis throughout the shoot. The examination of stomata by microscopy may be a useful screen for the directed isolation of additional cytokinesis mutations that are not embryo or seedling lethal     

Epigenetic silencing of Notch signaling in gastrointestinal cancers

The Notch signaling pathway drives proliferation, differentiation, apoptosis, cell fate choices and maintenance of stem cells during embryogenesis and in self-renewing tissues of the adult. In addition, aberrant Notch signaling has been implicated in several tumors, where Notch can function both as an oncogene or a tumor-suppressor gene, depending on the context.

This Extra View aims to review what is currently known about Notch signaling, in particular in gastrointestinal tumors, providing a summary of our data on Notch1 signaling in gastric cancer with results obtained in colorectal cancer (CRC).

We have already reported that the epigenetic regulation of the Notch ligand DLL1 controls Notch1 signaling activation in gastric cancer, and that Notch1 inhibition is associated with the diffuse type of gastric cancer. Here, we describe additional data showing that in CRC cell lines, unlike gastric cancer, DLL1 expression is not regulated by promoter methylation. Moreover, in CRC, Notch1 receptor is not affected by any mutation. These data suggest a different regulation of Notch1 signaling between gastric cancer and CRC.     

Matrine promotes apoptosis in SW480 colorectal cancer cells via elevating MIEF1-related mitochondrial division in a manner dependent on LATS2-Hippo pathway

Matrine, an alkaloid compound isolated from Sophora flavescens Ait, has been shown to exert cancer-killing actions in a variety of tumors; however, its anticancer mechanism in colorectal cancer (CRC) is not clear. The goal of our study was to characterize the anticancer effects and molecular mechanisms of matrine in SW480 CRC cells in vitro. Matrine treatment reduced mitochondrial metabolic function and ATP levels, repressed mitochondrial membrane potential, evoked mitochondrial reactive oxygen species accumulation, and promoted cyt-c-related mitochondrial apoptosis activation. In addition, we found that matrine treatment activated mitochondrial fission through upregulating mitochondrial elongation factor 1 (MIEF1); silencing of MIEF1 prevented matrine-mediated mitochondrial damage and reversed the decrease in SW480 cell viability. Moreover, matrine treatment affected MIEF1 expression via the large tumor suppressor-2 (LATS2)-Hippo axis, and LATS2 deficiency suppressed the anticancer actions exerted by matrine on SW480 cancer cells. In summary, we show for the first time that matrine inhibits SW480 cell survival by activating MIEF1-related mitochondrial division via the LATS2-Hippo pathway. These findings explain the anticancer mechanisms of matrine in CRC and also identify the LATS2-MIEF1 signaling pathway as an effective target for the treatment of CRC.     

Aberrant cell plate formation in the Arabidopsis thaliana microtubule organization 1 mutant

MICROTUBULE ORGANIZATION 1 encodes a microtubule-associated protein in Arabidopsis thaliana but different alleles have contradictory phenotypes. The original mutant mor1 alleles were reported to have disrupted cortical microtubules, swollen organs and normal cytokinesis, whereas other alleles, embryo-lethal gemini pollen 1 (gem1), have defective pollen cytokinesis. To determine whether MOR1 functions generally in cytokinesis, we examined the ultrastructure of cell division in roots of the original mor1-1 allele. Cell plates are misaligned, branched and meandering; the forming cell plates remain partly vesicular, with electron-dense or lamellar content. Phragmoplast microtubules are abundant but organized aberrantly. Thus, MOR1 functions in both phragmoplast and cortical arrays.     

Overexpression of IAA1 with domain II mutation impairs cell elongation and cell division in inflorescences and leaves of Arabidopsis

The auxin/indoleacetic acid (Aux/IAA) proteins are negative regulators of the auxin response factors (ARFs) that regulate expression of auxin-responsive genes. The Aux/IAA proteins have four conserved domains. Domain II is responsible for the rapid degradation of these proteins. Degradation of the Aux/IAA proteins, mediated by a SCF(TIR1) E3 ubiquitin protein ligase complex, is critical for auxin-regulated gene expression. Using a steroid-hormone-inducible system, we had previously shown that a protein-stability-enhancing mutation in domain II of IAA1 (iaa1) impaired diverse auxin responses. Inhibition of hypocotyl elongation, leaf expansion, and stem elongation by overexpression of iaa1 suggested that cell enlargement and/or cell division might be affected. We here examined the effects of the domain II mutation on cellular anatomy using light microscopy. Our results show that overexpression of iaa1 in Arabidopsis significantly reduced cell length and cell number and affected cell shape in inflorescences and leaves in a dexamethasone (DEX)-dependent manner. These results suggest that IAA1 might be involved in cell elongation as well as in cell division in the aerial parts of Arabidopsis plants. In addition, the formation of both phloem and xylem in leaves and stems was also impaired in a DEX-dependent manner, indicating a potential involvement of IAA1 in vascular development.     

Transposon-mediated mutations in the untranslated leader of maize Adh1 that increase and decrease pollen-specific gene expression.

The unstable mutation Adh1-Fm335 contains a Dissociation (Ds1) transposable element at position +53 in the untranslated leader of the maize Alcohol dehydrogenase-1 (Adh1) gene. Excision of Ds1 is known to generate new alleles with small additions and rearrangements of Adh1 DNA. We characterized 16 revertant alleles with respect to ADH1 activity levels in scutellum (nutritive tissue of the seed), anaerobic root, and pollen. Whereas gene expression was not different from the wild type in the sporophytic tissues of the scutellum and anaerobic root, there were strong allelic differences in pollen. One allele underexpressed pollen ADH1 at 48% of the wild-type level, and another overexpressed pollen ADH1 at 163% of the wild-type level. Quantitative RNase protection assays demonstrated that the mutant phenotypes reflected changes in the levels of steady state mRNA in pollen. These data provide a definitive demonstration of an overexpression mutant in plants and further show that marked increases in mRNA levels can follow minor alterations in central untranslated leader sequences. The nucleotide sequence of 12 new revertant alleles and the molecular mechanisms responsible for pollen-specific gene expression are discussed.     

The role of OsBRI1 and its homologous genes, OsBRL1 and OsBRL3, in rice

Since first identifying two alleles of a rice (Oryza sativa) brassinosteroid (BR)-insensitive mutant, d61, that were also defective in an orthologous gene in Arabidopsis (Arabidopsis thaliana) BRASSINOSTEROID INSENSITIVE1 (BRI1), we have isolated eight additional alleles, including null mutations, of the rice BRI1 gene OsBRI1. The most severe mutant, d61-4, exhibited severe dwarfism and twisted leaves, although pattern formation and differentiation were normal. This severe shoot phenotype was caused mainly by a defect in cell elongation and the disturbance of cell division after the determination of cell fate. In contrast to its severe shoot phenotype, the d61-4 mutant had a mild root phenotype. Concomitantly, the accumulation of castasterone, the active BR in rice, was up to 30-fold greater in the shoots, while only 1.5-fold greater in the roots. The homologous genes for OsBRI1, OsBRL1 and OsBRL3, were highly expressed in roots but weakly expressed in shoots, and their expression was higher in d61-4 than in the wild type. Based on these observations, we conclude that OsBRI1 is not essential for pattern formation or organ initiation, but is involved in organ development through controlling cell division and elongation. In addition, OsBRL1 and OsBRL3 are at least partly involved in BR perception in the roots.