Colin Brownlee

Reactive oxygen species produced by NADPH oxidase regulate plant cell growth.

Cell expansion is a central process in plant morphogenesis, and the elongation of roots and root hairs is essential for uptake of minerals and water from the soil. Ca2+ influx from the extracellular store is required for (and sets the rates of) cell elongation in roots. Arabidopsis thaliana rhd2 mutants are defective in Ca2+ uptake and consequently cell expansion is compromised—rhd2 mutants have short root hairs and stunted roots. To determine the regulation of Ca2+ acquisition in growing root cells we show here that RHD2 is an NADPH oxidase, a protein that transfers electrons from NADPH to an electron acceptor leading to the formation of reactive oxygen species (ROS). We show that ROS accumulate in growing wild-type (WT) root hairs but their levels are markedly decreased in rhd2 mutants. Blocking the activity of the NADPH oxidase with diphenylene iodonium (DPI) inhibits ROS formation and phenocopies Rhd2-. Treatment of rhd2 roots with ROS partly suppresses the mutant phenotype and stimulates the activity of plasma membrane hyperpolarization-activated Ca2+ channels, the predominant root Ca2+ acquisition system. This indicates that NADPH oxidases control development by making ROS that regulate plant cell expansion through the activation of Ca2+ channels.

The Phaeodactylum genome reveals the evolutionary history of diatom genomes.

Diatoms are photosynthetic secondary endosymbionts found throughout marine and freshwater environments, and are believed to be responsible for around one-fifth of the primary productivity on Earth. The genome sequence of the marine centric diatom Thalassiosira pseudonana was recently reported, revealing a wealth of information about diatom biology. Here we report the complete genome sequence of the pennate diatom Phaeodactylum tricornutum and compare it with that of T. pseudonana to clarify evolutionary origins, functional significance and ubiquity of these features throughout diatoms. In spite of the fact that the pennate and centric lineages have only been diverging for 90 million years, their genome structures are dramatically different and a substantial fraction of genes (∼40%) are not shared by these representatives of the two lineages. Analysis of molecular divergence compared with yeasts and metazoans reveals rapid rates of gene diversification in diatoms. Contributing factors include selective gene family expansions, differential losses and gains of genes and introns, and differential mobilization of transposable elements. Most significantly, we document the presence of hundreds of genes from bacteria. More than 300 of these gene transfers are found in both diatoms, attesting to their ancient origins, and many are likely to provide novel possibilities for metabolite management and for perception of environmental signals. These findings go a long way towards explaining the incredible diversity and success of the diatoms in contemporary oceans.

Calcium at the crossroads of signaling

### Calcium Signals: A Central Paradigm in Stimulus–Response Coupling Cells must respond to an array of environmental and developmental cues. The signaling networks that have evolved to generate appropriate cellular responses are varied and are normally composed of elements that include a

Communicating with Calcium

### Calcium as a Ubiquitous Signal in Plants All living cells use a network of signal transduction pathways to conduct developmental programs, obtain nutrients, control their metabolism, and cope with their environment. A major challenge for cell biologists is to understand the “language” of

The Ectocarpus genome and the independent evolution of multicellularity in brown algae

Brown algae (Phaeophyceae) are complex photosynthetic organisms with a very different evolutionary history to green plants, to which they are only distantly related. These seaweeds are the dominant species in rocky coastal ecosystems and they exhibit many interesting adaptations to these, often harsh, environments. Brown algae are also one of only a small number of eukaryotic lineages that have evolved complex multicellularity (Fig. 1). We report the 214 million base pair (Mbp) genome sequence of the filamentous seaweed Ectocarpus siliculosus (Dillwyn) Lyngbye, a model organism for brown algae, closely related to the kelps (Fig. 1). Genome features such as the presence of an extended set of light-harvesting and pigment biosynthesis genes and new metabolic processes such as halide metabolism help explain the ability of this organism to cope with the highly variable tidal environment. The evolution of multicellularity in this lineage is correlated with the presence of a rich array of signal transduction genes. Of particular interest is the presence of a family of receptor kinases, as the independent evolution of related molecules has been linked with the emergence of multicellularity in both the animal and green plant lineages. The Ectocarpus genome sequence represents an important step towards developing this organism as a model species, providing the possibility to combine genomic and genetic approaches to explore these and other aspects of brown algal biology further.

Exocytosis and endocytosis

Exocytosis is a general term used to denote vesicle fusion at the plasma membrane, and it is the final step in the secretory pathway that typically begins in the endoplasmic reticulum (ER), passes through the Golgi apparatus, and ends at the outside of the cell. Endocytosis refers to the recovery of

Inositol hexakisphosphate mobilizes an endomembrane store of calcium in guard cells

myo-Inositol hexakisphosphate (InsP6) is the most abundant inositol phosphate in cells, yet it remains the most enigmatic of this class of signaling molecule. InsP6 plays a role in the processes by which the drought stress hormone abscisic acid (ABA) induces stomatal closure, conserving water and ensuring plant survival. Previous work has shown that InsP6 levels in guard cells are elevated in response to ABA, and InsP6 inactivates the plasma membrane inward K+ conductance (IK,in) in a cytosolic calcium-dependent manner. The use of laser-scanning confocal microscopy in dye-loaded patch-clamped guard cell protoplasts shows that release of InsP6 from a caged precursor mobilizes calcium. Measurement of calcium (barium) currents ICa in patch-clamped protoplasts in whole cell mode shows that InsP6 has no effect on the calcium-permeable channels in the plasma membrane activated by ABA. The InsP6-mediated inhibition of IK,in can also be observed in the absence of external calcium. Thus the InsP6-induced increase in cytoplasmic calcium does not result from calcium influx but must arise from InsP6-triggered release of calcium from endomembrane stores. Measurements of vacuolar currents in patch-clamped isolated vacuoles in whole-vacuole mode showed that InsP6 activates both the fast and slow conductances of the guard cell vacuole. These data define InsP6 as an endomembrane-acting calcium-release signal in guard cells; the vacuole may contribute to InsP6-triggered Ca2+ release, but other endomembranes may also be involved.

Cell Fate Determination by the Cell Wall in Early Fucus Development

In multicellular plants, development starts with an asymmetric division of the zygote into two differentiated cells. The nature and distribution of fate-determining factors operating during embryogenesis remain largely obscure. Laser microsurgery was used here to dissect two-celled embryos of the alga Fucus spiralis. Removal of protoplasts from the cell wall induced dedifferentiation. However, isolated cells within the walls followed their restricted fate. Moreover, contact of one cell type with the isolated cell wall of the other cell type caused its fate to be switched. The cell wall thus appears to maintain the differentiated state and to direct cell fate in plant development.

Annexins: multifunctional components of growth and adaptation

Plant annexins are ubiquitous, soluble proteins capable of Ca(2+)-dependent and Ca(2+)-independent binding to endomembranes and the plasma membrane. Some members of this multigene family are capable of binding to F-actin, hydrolysing ATP and GTP, acting as peroxidases or cation channels. These multifunctional proteins are distributed throughout the plant and throughout the life cycle. Their expression and intracellular localization are under developmental and environmental control. The in vitro properties of annexins and their known, dynamic distribution patterns suggest that they could be central regulators or effectors of plant growth and stress signalling. Potentially, they could operate in signalling pathways involving cytosolic free calcium and reactive oxygen species.

The role of seawater endocytosis in the biomineralization process in calcareous foraminifera

Foraminifera are unicellular organisms that inhabit the oceans in various ecosystems. The majority of the foraminifera precipitate calcitic shells and are among the major CaCO3 producers in the oceans. They comprise an important component of the global carbon cycle and also provide valuable paleoceanographic information based on the relative abundance of stable isotopes and trace elements (proxies) in their shells. Understanding the biomineralization processes in foraminifera is important for predicting their calcification response to ocean acidification and for reliable interpretation of the paleoceanographic proxies. Most models of biomineralization invoke the involvement of membrane ion transporters (channels and pumps) in the delivery of Ca2+ and other ions to the calcification site. Here we show, in contrast, that in the benthic foraminiferan Amphistegina lobifera, (a shallow water species), transport of seawater via fluid phase endocytosis may account for most of the ions supplied to the calcification site. During their intracellular passage the seawater vacuoles undergo alkalization that elevates the CO32− concentration and further enhances their calcifying potential. This mechanism of biomineralization may explain why many calcareous foraminifera can be good recorders of paleoceanographic conditions. It may also explain the sensitivity to ocean acidification that was observed in several planktonic and benthic species.