In Vivo Functions of an ER Ca2+/Mn2+ Pump

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Although Ca²⁺ and Mn²⁺ are essential plant nutrients, they are potentially toxic at high external concentrations. Both cations enter plant cells down an electrochemical gradient. In the cytoplasm, their levels are tightly regulated in the range of 0.1 to 0.2 µm. Cytosolic Ca²⁺ is maintained at low levels by ATP-driven pumps and Ca²⁺/H⁺ antiporters located at membranes, including the plasma membrane, vacuole, and endoplasmic reticulum (ER). Mn²⁺ is believed to be accumulated mostly in the vacuole and chloroplast. Arabidopsis contains 15 putative Ca²⁺-ATPases, as predicted from the completed genome sequence, though the in vivo function of each pump is unknown. One of the best characterized of these Ca²⁺-ATPases is ECA1, an ER-type Ca²⁺ pump. In this issue, Wu et al. (pp. 128-137) show that ECA1 has a dual role in both Ca²⁺ and Mn²⁺ homeostasis. They provide biochemical evidence that ECA1 provides approximately 70% of the total ER-type Ca²⁺ pump activity in Arabidopsis. Surprisingly, a plant with a T-DNA disruption (eca1-1) of this Ca²⁺-ATPase exhibits a wild-type phenotype when grown under standard nutrient conditions. Under conditions of Ca²⁺ deprivation, however, the growth of the mutant plants is impaired, demonstrating that ECA1 provides an important function in Ca²⁺ nutrition. The authors also provide biochemical and genetic evidence that ECA1 serves as an Mn²⁺ pump, and that it confers tolerance to toxic levels of Mn²⁺. Under conditions of high Mn²⁺, the root hairs of the mutants failed to elongate, suggesting impairment in tip growth processes. These studies provide the first genetic evidence for the in vivo function of a Ca²⁺ pump in plants.

	Novel Cytokinesis Mutants of Arabidopsis

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The cells of plant cytokinesis mutants are typically enlarged, have incomplete cell walls, and are either multinucleate or contain enlarged polyploid nuclei or both. In this issue, Müller et al. (pp. 312-324) report on their identification of two new loci in Arabidopsis, PLEIADE (PLE) and HYADE (HYA), whose mutant phenotypes exhibit typical features of cytokinesis-defective mutants, but only in root cells. The ple and hya mutants range from having very thick, short roots to elongated roots with a wavy growth pattern and enhanced lateral root initiation.

Visualization of the nuclei in mutants revealed that the giant root cells contain up to 32 nuclei, indicating that these cells undergo karyokinesis without cytokinesis (Fig. 1). The punctate appearance of this multinucleate phenotype is the reason for naming the genes after the stellar constellations, the Pleiades and Hyades (Fig. 2). The ple and hya mutants contain partially formed transverse cell walls. During cell division, these multinucleate cells divide synchronously and influence the position of microtubule arrays including the preprophase band, the mitotic spindle, and the phragmoplast. The enhanced phenotypes of ple/hya double mutants point to a role of PLE and HYA in the same process. In contrast to the single mutants, the double mutants were not viable on soil and were not fertile. The strong root phenotypes of the ple and hya alleles indicate that the genes may encode organ-specific components needed primarily during root development. The authors propose that a certain threshold activity of the PLE and HYA gene products is needed to stabilize cytokinetic structures.

View larger version (111K): [in this window] [in a new window]   Figure 1.   The pleiade mutant of Arabidopsis exhibits defects in cytokinesis in root cells only. Note the enlarged, irregular, multinucleate cells.

View larger version (91K): [in this window] [in a new window]   Figure 2.   The Pleiades constellation was the inspiration for naming the pleiade mutant of Arabidopsis. ©Anglo-Australian Observatory/Royal Observatory, Edinburgh.

	Plant-Like Ethylene Receptor Genes in Cyanobacteria

TOP In Vivo Functions of... Novel Cytokinesis Mutants of... Plant-Like Ethylene Receptor... Regulated Expression of... Engineering Herbicide...

The ethylene receptors of higher plants are similar to the two-component signaling proteins that are widely used by bacteria in responding to diverse stimuli. In this issue, Mount and Chang (pp. 10-14) report that two genes in the newly completed genome sequence of the cyanobacterium Anabaena sp. strain PCC 7120 encode homologs of plant ethylene receptors. Moreover, a re-analysis of a previously reported ethylene-binding protein from the cyanobacterium Synechocystis sp. reveals it to be another example of an ethylene receptor homolog. The basic structure of plant ethylene receptors consists of an amino-terminal ethylene-binding domain linked to a two-component His kinase domain by a GAF domain. The predicted Anabaena proteins have approximately 40% amino acid identity with plant ethylene receptors in the ethylene-binding domain and 26% to 35% identity in the His kinase domain. Similar to certain subtypes of plant ethylene receptors, one of the Anabaena homologs has a carboxyl-terminal receiver domain, which shares up to 32% amino acid identity with those of Arabidopsis ethylene receptors, including a conserved phosphorylation site. With the exception of these two cyanobacterial genomes, no ethylene receptor sequences are found in any of the 70 fully sequenced microbial genomes, but the authors did detect three additional examples of ethylene-binding domain sequences in unfinished sequences from three highly divergent bacterial species other than cyanobacteria. The observed distribution of ethylene receptor sequences is most consistent with a cyanobacterial (plastid) origin of plant ethylene receptors and rare horizontal transfer of ethylene receptor genes from cyanobacteria into diverse bacterial lineages.

	Regulated Expression of Arabidopsis Phosphate Transporters

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Many plants show an enhanced capacity for phosphate uptake in response to P deficiency. This increase has been correlated with an increased number of high-affinity phosphate transporters in the plasma membrane. The protein products of two Arabidopsis genes (AtPT1 and AtPT2) have been shown to function as high-affinity phosphate transporters that serve to translocate phosphate from P-deficient media into the cytoplasm. Karthikeyan et al. (pp. 221-233) have extensively analyzed the transcriptional and spatial regulation of low-phosphate-induced gene expression using three reporter genes regulated by the AtPT1 and AtPT2 transporter promoters. Activation of the genes was rapid, repressible, and specific in response to changes in phosphate availability. The phytohormones auxin and cytokinin suppressed the expression of a reporter gene driven by the AtPT1 promoter, suggesting that hormones may be involved in regulation of some component(s) of the P starvation response pathway. There were distinct differences in the patterns of expression of reporter genes driven by the AtPT1 and AtPT2 promoters in roots of P-starved plants. For example, AtPT1 promoter-driven reporter gene activity was lacking in root tips. In contrast, AtPT2 promoter-driven reporter gene expression was found all along the root and was particularly strong in the vascular region, indicating that it may also be involved in phosphate transfer into the vascular tissues. Both AtPT1 and AtPT2 promoters strongly activated reporter gene expression in elongating root hairs, which are known to be major players in the acquisition of phosphate. Evidence is also presented for a potential role of high-affinity phosphate transporters in mobilizing phosphate into reproductive organs. The results suggest that members of the phosphate transporter family have similar but non-redundant functions in plants.

	Engineering Herbicide Metabolism Using Cytochrome P450

TOP In Vivo Functions of... Novel Cytokinesis Mutants of... Plant-Like Ethylene Receptor... Regulated Expression of... Engineering Herbicide...

The genetic engineering of crops for enhanced herbicide metabolism represents a good potential strategy for increasing herbicide tolerance. This is because the phytotoxic compound is chemically altered and there is no interference with primary metabolism and no residual herbicide remains in the plant. In vivo and in vitro experimentation has demonstrated the involvement of cytochromes P450 in the metabolism of all major classes of herbicides and their contribution to both herbicide selectivity and weed resistance. The cytochrome P450 monooxygenases, which constitute the largest family of enzymatic proteins in higher plants (272 cytochrome P450 genes have been found in Arabidopsis), offer a wide potential source of herbicide-detoxifying proteins. In this issue, Didierjean et al. (pp. 179-189) report that increased herbicide metabolism and tolerance in tobacco (Nicotiana tabacum) and Arabidopsis is achieved by the ectopic constitutive expression of a specific Helianthus tuberosus xenobiotic-inducible cytochrome P450 (CYP76B1). CYP76B1 catalyzes the rapid oxidative dealkylation of various phenylurea herbicides to yield non-phytotoxic metabolites. Transformation with CYP76B1 conferred on both tobacco and Arabidopsis a 20-fold increase in tolerance to linuron, a compound detoxified by a single dealkylation, and a 10-fold increase in tolerance to isoproturon or chlortoluron, which need successive catalytic steps for detoxification. Aside from increased herbicide tolerance, the ectopic expression of CYP76B1 has no other visible phenotypic effects on the transgenic plants. The data also indicate that CYP76B1 can function as a selectable marker for plant transformation, allowing efficient selection both in vitro and on soil-grown plants. Plants expressing CYP76B1 may also be useful in the phytoremediation of contaminated sites.