Sherryl R. Bisgrove

The Microtubule Plus-End Binding Protein EB1 Functions in Root Responses to Touch and Gravity Signals in Arabidopsis

Microtubules function in concert with associated proteins that modify microtubule behavior and/or transmit signals that effect changes in growth. To better understand how microtubules and their associated proteins influence growth, we analyzed one family of microtubule-associated proteins, the END BINDING1 (EB1) proteins, in Arabidopsis thaliana (EB1a, EB1b, and EB1c). We find that antibodies directed against EB1 proteins colocalize with microtubules in roots, an observation that confirms previous reports using EB1-GFP fusions. We also find that T-DNA insertion mutants with reduced expression from EB1 genes have roots that deviate toward the left on vertical or inclined plates. Mutant roots also exhibit extended horizontal growth before they bend downward after tracking around an obstacle or after a 90° clockwise reorientation of the root. These observations suggest that leftward deviations in root growth may be the result of delayed responses to touch and/or gravity signals. Root lengths and widths are normal, indicating that the delay in bend formation is not due to changes in the overall rate of growth. In addition, the genotype with the most severe defects responds to low doses of microtubule inhibitors in a manner indistinguishable from the wild type, indicating that microtubule integrity is not a major contributor to the leftward deviations in mutant root growth.

TIPs and Microtubule Regulation. The Beginning of the Plus End in Plants

Plants have evolved novel microtubule (MT) arrays to regulate cell division and cell expansion. How these MT arrangements are managed has been a question of long-standing interest to plant cell biologists. Do plants have unique ways of regulating MTs or have they co-opted mechanisms that are

Asymmetric Cell Divisions: Zygotes of Fucoid Algae as a Model System

Asymmetric cell divisions are commonly used across diverse phyla to generate different kindsof cells during development. Although asymmetric divisions play important roles during developmentin plants, algae, fungi, and animals, emerging data indicate that there is some variability amongstthe mechanisms that are at play in these different organisms. Zygotes of fucoid algae have long servedas models for understanding early developmental processes including cell polarization and asymmetriccell division. In addition, brown algae are phylogenetically distant from other organisms, includingplant models, a feature that makes them interesting from a comparative perspective (Andersen2004; Peters et al. 2004). This monograph focuses on advances made toward understanding howasymmetric divisions are regulated in fucoid algae and, where appropriate, comparisons are made tohigher plant zygotes.

The microtubule associated protein END BINDING 1 represses root responses to mechanical cues.

The ability of roots to navigate around rocks and other debris as they grow through the soil requires a mechanism for detecting and responding to input from both touch and gravity sensing systems. The microtubule associated protein END BINDING 1b (EB1b) is involved in this process as mutants have defects responding to combinations of touch and gravity cues. This study investigates the role of EB1b in root responses to mechanical cues. We find that eb1b-1 mutant roots exhibit an increase over wild type in their response to touch and that the expression of EB1b genes in transgenic mutants restores the response to wild type levels, indicating that EB1b is an inhibitor of the response. Mutant roots are also hypersensitive to increased levels of mechanical stimulation, revealing the presence of another process that activates the response. These findings are supported by analyses of double mutants between eb1b-1 and seedlings carrying mutations in PHOSPHOGLUCOMUTASE (PGM), ALTERED RESPONSE TO GRAVITY1 (ARG1), or TOUCH3 (TCH3), genes that encode proteins involved in gravity sensing, signaling, or touch responses, respectively. A model is proposed in which root responses to mechanical cues are modulated by at least two competing regulatory processes, one that promotes touch-mediated growth and another, regulated by EB1b, which dampens root responses to touch and enhances gravitropism.

Microtubule Plus End-Tracking Proteins and Their Activities in Plants

Microtubules form a dynamic system of filaments that have essential roles in all eukaryotic cells. Although microtubules are well-conserved across eukaryotic phyla, the organization and function of the microtubule cytoskeleton has evolved in ways that set higher plants apart from other organisms. The mechanisms that underlie these differences are topics of long-standing interest to plant cell biologists. A feature that is a key to microtubule function is their dynamic nature; they usually exist in states of growth or shrinkage. Their ability to grow, depolymerize, and then re-grow in new directions allows them to explore the cytoplasm and to rearrange into different configurations as needed. Microtubules also function in concert with a fleet of proteins that regulate microtubule activities within the cell. This chapter focuses on a specialized group of microtubule associated proteins (MAPs) that localize to the more active or plus ends of microtubules. From their position at the active end of the microtubule, these plus end-tracking proteins, or +TIPs, have a large influence on microtubule behaviour. They regulate growth and shrinkage rates and mediate interactions of the microtubule end with other proteins or structures in the cell. A number of +TIPs have been identified from several organisms including plants. The proteins form a structurally and functionally diverse group that have very little in common aside from an affinity for microtubule plus ends. Current models propose that these proteins form a dynamic +TIP network that is remodelled to include different sets of proteins depending on the needs of the cell. Here we discuss the repertoire of +TIP families found in plants. Plant cells have homologs corresponding to several proteins with plus-end binding activity in other organisms. Analyses indicate that there is some functional conservation amongst these proteins, although there are also cases where the plant proteins have been modified to function in different ways. In addition to conserved proteins, plants use at least one +TIP family that is not found in other eukaryotic genomes and there are some families that appear to be absent in higher plant lineages. The research to date suggests that plants have a somewhat modified +TIP network which is uniquely suited to the needs of the plant cell.

The microtubule-associated protein END BINDING1b, auxin, and root responses to mechanical cues

The ability of roots to penetrate through the soil and maneuver around rocks and other impenetrable objects requires a system for modulating output from mechanosensory response networks. The microtubule-associated protein END BINDING1b (EB1b) has a role in this process; it represses root responses to mechanical cues. In this study, a possible relationship between EB1b and auxin during root responses to mechanical cues was investigated. We found that eb1b-1-mutant roots are more sensitive than wild-type roots to chemicals that disrupt auxin transport, whereas the roots of mutants with defects in auxin transport are resistant to these treatments. Using seedlings that express the auxin-sensitive DR5rev::GFP construct, we also found that wild-type and eb1b-1 roots treated with the auxin transport inhibitor naphthylphthalamic acid exhibited dose-dependent reductions in basipetal auxin transport that were indistinguishable from each other. The responses of eb1b-1 roots to mechanical cues were also enhanced over wild type in the presence of p-chlorophenoxyisobutyric acid, a chemical thought to inhibit auxin signaling. Finally, roots of eb1b-1 and wild-type plants exhibited slight increases in loop formation in response to increasing levels of exogenously applied indole-3-acetic acid or 1-naphthalene acetic acid. Taken together, these results suggest that the repression of loop formation by EB1b and auxin transport/signaling occurs by different mechanisms.

Identification of genes that contribute to drought stress tolerance in Populus

Background The cultivation of poplars (Populus spp.) is favored for forestry and reclamation purposes all over the northern hemisphere where they represent a commercially important resource. Poplars may become a component of programs to optimize carbon sequestration however; poplars are generally regarded as drought sensitive. The patterns of episodic drought over the last decade suggest that the development of drought tolerant poplar genotypes could be a useful tool to achieve sustained forest productivity [1]. Previous reports have shown that expression of hundreds of poplar genes changes in response to drought, presenting a problem in the identification of genes that are more important than others in counteracting the harmful effects of drought [2,3]. The genus Populus contains many fast growing hybrids that show varied drought tolerance according to genotype [4]. Hence, there is genetic variation among poplar hybrids that can be used to identify genes that contribute to drought stress tolerance. Despite extensive physiological and morphological descriptions of the response of Populus to drought, little work has been undertaken to explain genotype differences at the gene level. Therefore, this research has been undertaken and its major objective is to identify the genes that contribute to drought stress tolerance in poplar by correlating the physiological responses to gene expression. These genes may potentially be used as molecular markers in the drought tolerance breeding programs.