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Molecular analysis of the SCARECROW gene in maize reveals a common basis for radial patterning in diverse meristems.

Maize and Arabidopsis root apical meristems differ in several aspects of their radial organization and ontogeny. Despite the large evolutionary distance and differences in root radial patterning, analysis of the putative maize ortholog of the Arabidopsis patterning gene SCARECROW (SCR) revealed expression localized to the endodermis, which is similar to its expression in Arabidopsis. Expression in maize extends through the quiescent center, a population of mitotically inactive cells formerly thought to be undifferentiated and to lack radial pattern information. Zea mays SCARECROW (ZmSCR), the putative maize SCR ortholog, was used as a molecular marker to investigate radial patterning during regeneration of the root tip after either whole or partial excision. Analysis of the dynamic expression pattern of ZmSCR as well as other markers indicates the involvement of positional information as a primary determinant in regeneration of the root radial pattern.

Amino Acid Sequence↗

Cell wall invertase expression at the apical meristem alters floral, architectural, and reproductive traits in Arabidopsis thaliana.

Resource allocation is a major determinant of plant fitness and is influenced by external as well as internal stimuli. We have investigated the effect of cell wall invertase activity on the transition from vegetative to reproductive growth, inflorescence architecture, and reproductive output, i.e. seed production, in the model plant Arabidopsis thaliana by expressing a cell wall invertase under a meristem-specific promoter. Increased cell wall invertase activity causes accelerated flowering and an increase in seed yield by nearly 30%. This increase is caused by an elevation of the number of siliques, which results from enhanced branching of the inflorescence. On the contrary, as cytosolic enzyme, the invertase causes delayed flowering, reduced seed yield, and branching. This demonstrates that invertases not only are important in determining sink strength of storage organs but also play a role in regulating developmental processes.

Arabidopsis↗

Time-lapse analysis of stem-cell divisions in the Arabidopsis thaliana root meristem.

In the Arabidopsis root, asymmetric stem-cell divisions produce daughters that form the different root cell types. Here we report the establishment of a confocal tracking system that allows the analysis of numbers and orientations of cell divisions in root stem cells. The system provides direct evidence that stem cells have lower division rates than cells in the proximal meristem. It also allows tracking of cell division timing, which we have used to analyse the synchronization of root cap divisions. Finally, it gives new insights into lateral root cap formation: epidermal stem-cell daughters can rotate the orientation of the division plane like the stem cell.

Arabidopsis↗

Molecular characterization of the duplicated meristem identity genes HvAP1a and HvAP1b in barley.

The vernalization gene VRN-1 has been identified as a MADS-box transcription factor orthologous to the meristem identity gene APETALA1 (AP1). A single copy of this gene was found in diploid wheat, but 2 copies were reported in barley. In this study, we present a detailed characterization of these 2 copies to understand their respective roles in the vernalization response. We identified 2 groups of barley bacterial artificial chromosomes (BACs), each containing 1 AP1 copy designated hereafter as HvAP1a and HvAP1b. A physical map of the VRN-H1 region showed that the HvAP1a BACs were part of the VRN-H1 region but that the HvAP1b BACs were not. Numerous structural changes were observed between the barley and wheat VRN-1 physical maps. In a population segregating for VRN-H1, the HvAP1a gene cosegregated with growth habit, suggesting that HvAP1a is the barley vernalization gene VRN-H1. The other copy, HvAP1b, was mapped on the centromeric region of chromosome 1H, the chromosome where vernalization gene VRN-H3 was previously mapped. We developed a mapping population segregating for VRN-H3 and showed that 2 molecular makers flanking HvAP1b locus were not linked to growth habit. The HvAP1b copy has a complete deletion of the first 2 exons, suggesting that it is a truncated pseudogene and not a candidate for VRN-H3. In summary, this study contributed a detailed physical map of the barley VRN-H1 region, showed several structural differences with the orthologous wheat region, and clarified the identity of the barley VRN-H1 gene.

Base Sequence↗

Leaving the meristem behind: regulation of KNOX genes.

The mechanism by which the plant reserves some cells as pluripotent stem cells while partitioning others into differentiated leaf tissue is fundamental to plant development. New work in Arabidopsis elucidates the genetic circuitry that distinguishes meristem from leaf.

Arabidopsis↗

Mutations in Arabidopsis condensin genes disrupt embryogenesis, meristem organization and segregation of homologous chromosomes during meiosis.

Proper chromatin condensation and sister chromatid resolution are essential for the maintenance of chromosomal integrity during cell division, and is in part mediated by a conserved multisubunit apparatus termed the condensin complex. The core subunits of the complex are members of the SMC2 (Structural Maintenance of Chromosomes) and SMC4 gene families. We have cloned an Arabidopsis gene, AtCAP-E1, which is a functional ortholog of the yeast SMC2 gene. A second, highly homologous SMC2 gene, AtCAPE-2, was identified by the Arabidopsis genome project. SMC2 gene expression in Arabidopsis was correlated with the mitotic activity of tissues, with high level expression observed in meristematic cells. The two genes are differentially expressed with AtCAP-E1 accounting for more than 85% of the total SMC2 transcript pool. The titan3 mutant is the result of a T-DNA insertion into AtCAP-E1, but other than subtle endosperm defects, titan3 is viable and fecund. We identified a T-DNA insertion mutant of AtCAP-E2, which showed no obvious mutant phenotype, indicating that the two genes are functionally redundant. Genetic crosses were employed to examine the consequences of reduced SMC2 levels. Both male and female gametogenesis were compromised in double mutant spores. Embryo lethality was observed for both double homozygous and AtCAP-E1(-/-), AtCAP-E2(+/-) plants; arrest occurred at or before the globular stage and was associated with altered planes of cell division in both the suspensor and the embryo. Down regulation of both genes by antisense technology, as well as in AtCAP-E1(+/-), AtCAP-E2(-/-) plants results in meristem disorganization and fasciation. Our data are consistent with the interpretation that threshold levels of SMC2 proteins are required for normal development and that AtCAP-E2 may have a higher affinity for its target than AtCAP-E1.

Adenosine Triphosphatases↗

MicroRNA regulation of the CUC genes is required for boundary size control in Arabidopsis meristems.

We have analysed the role of a microRNA, miR164, in boundary formation during organ initiation from Arabidopsis meristems. The establishment and maintenance of the boundary domain are controlled by three partially redundant genes, CUP-SHAPED COTYLEDON1 (CUC1), CUC2 and CUC3. We show that miR164 overexpression phenocopies the cuc1 cuc2 double mutant by inducing post-transcriptional downregulation of CUC1 and CUC2 but not CUC3 mRNA levels. Disruption of CUC2 regulation by miR164, either by making CUC2 resistant to the miRNA or by reducing miRNA levels leads to similar enlarged boundary domains. We relate this enlargement to the division patterns of the boundary cells. We propose that miR164 constrains the expansion of the boundary domain, by degrading CUC1 and CUC2 mRNAs.

Arabidopsis↗

Leaf polarity and meristem formation in Arabidopsis.

Shoot apical meristems (SAMs) of seed plants are small groups of pluripotent cells responsible for making leaves, stems and flowers. While the primary SAM forms during embryogenesis, new SAMs, called axillary SAMs, develop later on the body of the plant and give rise to branches. In Arabidopsis plants, axillary SAMs develop in close association with the adaxial leaf base at the junction of the leaf and stem (the leaf axil). We describe the phenotype caused by the Arabidopsis phabulosa-1d (phb-1d) mutation. phb-1d is a dominant mutation that causes altered leaf polarity such that adaxial characters develop in place of abaxial leaf characters. The adaxialized leaves fail to develop leaf blades. This supports a recently proposed model in which the juxtaposition of ad- and abaxial cell fates is required for blade outgrowth. In addition to the alteration in leaf polarity, phb-1d mutants develop ectopic SAMs on the undersides of their leaves. Also, the phb-1d mutation weakly suppresses the shoot meristemless (stm) mutant phenotype. These observations indicate an important role for adaxial cell fate in promoting the development of axiallary SAMs and suggest a cyclical model for shoot development: SAMs make leaves which in turn are responsible for generating new SAMs.

Arabidopsis↗

Initiation of shoot apical meristem in rice: characterization of four SHOOTLESS genes.

The regulatory mechanism of shoot apical meristem (SAM) initiation is an important subject in developmental plant biology. We characterized nine recessive mutations derived from four independent loci (SHL1-SHL4) causing the deletion of the SAM. Radicles were produced in these mutant embryos. Concomitant with the loss of SAM, two embryo-specific organs, coleoptile and epiblast, were lost, but the scutellum was formed normally. Therefore, differentiation of radicle and scutellum is regulated independently of SAM, but that of coleoptile and epiblast may depend on SAM. Regeneration experiments using adventitious shoots from the scutellum-derived calli showed that no adventitious shoots were regenerated in any shl mutant. However, small adventitious leaves were observed in both mutant and wild-type calli, but they soon became necrotic and showed no extensive growth. Thus, leaf primordia can initiate in the absence of SAM, but their extensive growth requires the SAM. An in situ hybridization experiment using a rice homeobox gene, OSH1, as a probe revealed that shl1 and shl2 modified the expression domain of OSH1, but normal expression of OSH1 was observed in shl3 and shl4 embryos. Accordingly, SHL1 and SHL2 function upstream of OSH1, and SHL3 and SHL4 downstream or independently of OSH1. These shl mutants are useful for elucidating the genetic program driving SAM initiation and for unraveling the interrelationships among various organs in grass embryos.

Gene Deletion↗

Redundant regulation of meristem identity and plant architecture by FRUITFULL, APETALA1 and CAULIFLOWER.

The transition from vegetative to reproductive phases during Arabidopsis development is the result of a complex interaction of environmental and endogenous factors. One of the key regulators of this transition is LEAFY (LFY), whose threshold levels of activity are proposed to mediate the initiation of flowers. The closely related APETALA1 (AP1) and CAULIFLOWER (CAL) meristem identity genes are also important for flower initiation, in part because of their roles in upregulating LFY expression. We have found that mutations in the FRUITFULL (FUL) MADS-box gene, when combined with mutations in AP1 and CAL, lead to a dramatic non-flowering phenotype in which plants continuously elaborate leafy shoots in place of flowers. We demonstrate that this phenotype is caused both by the lack of LFY upregulation and by the ectopic expression of the TERMINAL FLOWER1 (TFL1) gene. Our results suggest that the FUL, AP1 and CAL genes act redundantly to control inflorescence architecture by affecting the domains of LFY and TFL1 expression as well as the relative levels of their activities.

Arabidopsis↗

The ASYMMETRIC LEAVES2 gene of Arabidopsis thaliana regulates formation of a symmetric lamina, establishment of venation and repression of meristem-related homeobox genes in leaves.

The asymmetric leaves2 (as2) mutant of Arabidopsis thaliana generated leaf lobes and leaflet-like structures from the petioles of leaves in a bilaterally asymmetric manner. Both the delayed formation of the primary vein and the asymmetric formation of secondary veins were apparent in leaf primordia of as2 plants. A distinct midvein, which is the thickest vein and is located in the longitudinal center of the leaf lamina of wild-type plants, was often rudimentary even in mature as2 leaves. However, several parallel veins of very similar thickness were evident in such leaves. The complexity of venation patterns in all leaf-like organs of as2 plants was reduced. The malformed veins were visible before the development of asymmetry of the leaf lamina and were maintained in mature as2 leaves. In vitro culture on phytohormone-free medium of leaf sections from as2 mutants and from the asymmetric leaves1 (as1) mutant, which has a phenotype similar to that of as2, revealed an elevated potential in both cases for regeneration of shoots from leaf cells. Analysis by the reverse transcription-polymerase chain reaction showed that transcripts of the KNAT1, KNAT2 and KNAT6 (a recently identified member of the class 1 knox family) genes accumulated in the leaves of both as2 and as1 plants but not of wild type. Transcripts of the STM gene also accumulated in as1 leaves. These findings suggest that, in leaves, the AS2 and AS1 genes repress the expression of these homeobox genes, which are thought to maintain the indeterminate cell state in the shoot apical meristem. Taken together, our results suggest that AS2 and AS1 might be involved in establishment of a prominent midvein and of networks of other veins as well as in the formation of the symmetric leaf lamina, which might be related to repression of class 1 knox homeobox genes in leaves.

Arabidopsis↗

The CUP-SHAPED COTYLEDON1 gene of Arabidopsis regulates shoot apical meristem formation.

In higher plants, molecular mechanisms regulating shoot apical meristem (SAM) formation and organ separation are largely unknown. The CUC1 (CUP-SHAPED COTYLEDON1) and CUC2 are functionally redundant genes that are involved in these processes. We cloned the CUC1 gene by a map-based approach, and found that it encodes a NAC-domain protein highly homologous to CUC2. CUC1 mRNA was detected in the presumptive SAM during embryogenesis, and at the boundaries between floral organ primordia. Surprisingly, overexpression of CUC1 was sufficient to induce adventitious shoots on the adaxial surface of cotyledons. Expression analyses in the overexpressor and in loss-of-function mutants suggest that CUC1 acts upstream of the SHOOT MERISTEMLESS gene.

Amino Acid Sequence↗

Structure-activity relationship study of host-specific phytotoxins (AM-toxin analogs) using a new assay method with leaves from apple meristem culture.

AM-toxins are host-specific phytotoxins of the Alternaria alternata apple pathotype, which induce necrosis on apple leaves. In this study, we developed a new assay to measure the necrotic activity of AM-toxin analogs using cultured leaves from meristem cells. This method was not only more sensitive to AM-toxin I, but also more reliable than the previous one that used tree leaves due to the homogeneous nature of cultured leaves and to the method of application of toxins. Using this assay method we investigated a structure-activity relationship of AM-toxin analogs synthesized in this study. Most residues and the macrocyclic ring structure were strictly recognized by AM-toxin putative receptor, whereas the L-Ala binding subsite of the receptor allowed for side chain structures with various stereoelectronic properties. These findings are important for designing ligands for further experimental probing of the nature of the receptor.

Alternaria↗

Metaphase chromosome accumulation and flow karyotypes in rice (Oryza sativa L.) root tip meristem cells.

Highly efficient cell synchronization and metaphase chromosome accumulation in rice root tip cells were achieved. Flow cytometric analysis was performed for obtaining optimal parameters to synchronize the cell cycles. High mitotic indices (about 57.6% in root tip meristemic area) were obtained by treating seedlings with 0.5 cm length using 0.5 mM hydroxyurea at 30 degrees C for 4 h, incubating in a hydroxyurea-free solution for 30 min, and then treating with 0.3 microM trifluralin for 3 h. After trifluralin treatment, incubation in distilled water for 15 min reduced chromosome clumping on metaphase spread. Uniformity of seed germination at the time of treatment is a critical parameter for obtaining high metaphase index. Isolated rice chromosomes were suitable for flow cytometric analysis and chromosome sorting. The morphology of flow sorted metaphase chromosomes was intact.

Cell Cycle↗

A critical role of sterols in embryonic patterning and meristem programming revealed by the fackel mutants of Arabidopsis thaliana.

Here we report a novel Arabidopsis dwarf mutant, fackel-J79, whose adult morphology resembles that of brassinosteroid-deficient mutants but also displays distorted embryos, supernumerary cotyledons, multiple shoot meristems, and stunted roots. We cloned the FACKEL gene and found that it encodes a protein with sequence similarity to both the human sterol reductase family and yeast C-14 sterol reductase and is preferentially expressed in actively growing cells. Biochemical analysis indicates that the fk-J79 mutation results in deficient C-14 sterol reductase activity, abnormal sterol composition, and reduction of brassinosteroids (BRs). Unlike other BR-deficient mutants, the defect of hypocotyl elongation in fk-J79 cannot be corrected by exogenous BRs. The unique phenotypes and sterol composition in fk-J79 indicate crucial roles of sterol regulation and signaling in cell division and cell expansion in embryonic and post-embryonic development in plants.

Alleles↗

[Assessment of phyto- and cytotoxic effects of heavy metals and aluminum compounds using onion apical root meristem].

The effects of different concentrations (10(-6)-10(-3) M) of salts of six metals (cadmium, lead, nickel, aluminum, copper, and zinc) on the root growth of onion (Allium cepa L.) seedling and cell division, chromosome and nucleus morphology in root tip cells were studied. The obtained results showed that tested compounds inhibit the root growth, reduce the mitotic activity of meristem cells and cause the chromosome and nucleus irregularities. On the basis of these data two rows of metal salt toxicity were proposed--for effective concentrations (EC50) of toxic action: CuSO4 > CdCl2 > NiSO4 > Pb(CH3COO)2 > Al(NO3)3 > ZnSO4, and for sublethal and lethal effects of investigated substances: CuSO4 > Pb(CH3COO)2 > CdCl2 > ZnSO4 > NiSO4 > Al(NO3)3.

Aluminum↗

[Cytogenetic effects of toxic metal salts on apical meristem cells of Allium cepa L. seed roots].

Comparative analysis of the influence of the salts of Cd, Pb, Ni, Al, Cu and Zn at concentrations 10(-6)-10(-3) M on cytogenetic parameters of Allium cepa apical meristem cells was carried out. It was shown that these metal compounds could induce both the clastogenic and the aneugenic effects (including mitosis and cytokinesis disturbances). Based on ability to cause the chromosome aberrations the metal salts were arranged in such row of genotoxicity: ZnSO4 > Pb(CH3COO)2 > Al(NO3)3 approximately NiSO4 > CdCl2 > CuSO4. The investigated compounds of cadmium, lead, nickel and aluminum were found to be more effective aneugens than clastogens. According to our data NiSO4 had the strongest antimitotic activity (it increased the percentage of chromosome lagging of, multipolar anaphases, C-mitoses in 69.6 times, compared to the control). The ability of other metal salts to induce such abnormalities decreased in the range: CdCl2 > Al(NO3)3 > Pb(CH3COO)2 > CuSO4 > ZnSO4. Increasing of binuclear cells on the slides was revealed after treatment by all tested substances, however Ni sulfate was found as the strongest inducer of cytokinesis abnormalities. Presented results allowed to suggest that cytoskeleton could be a cell target for salts of some metals (especially for NiSO4).

Cell Division↗

[Aluminum induces chromosome aberrations in wheat root meristem cells].

The yield and pattern of chromosome structure aberrations in wheat seedlings treated with aluminum nitrate and aluminum sulfate at various concentrations have been determined by the anaphase method. Aluminum has a genotoxic effect causing genome, chromatid, and chromosome aberrations in apical root meristem cells. The relationship between the total yield of structural mutations and the aluminum concentration follows a bell-shaped curve. The mutagenic activity of aluminum nitrate peaks at 10(-3) mg/ml, which is twice as high as the permissible concentration limit (PCL) of aluminum in potable water. The maximum of the mutagenic activity of aluminum sulfate is observed at 5 x 10(-4) mg/ml, i.e., one PCL. Tap water boiled for 2 h in an aluminum vessel has virtually no genotoxic effect on wheat cells.

Aluminum↗