Using chemical genomics to study cell wall formation and cell growth in Arabidopsis thaliana and Penium margaritaceum.

The cell wall is directly involved in cell growth, and its ability to loosen and rearrange allows for cell expansion through the existing turgor pressure. Thus, information on cell wall deposition and rearrangement can provide insights into the overall plant growth. This chapter describes two methods that can be used to evaluate cell expansion (1) in the model plant Arabidopsis thaliana and (2) the model alga Penium margaritaceum. These methods are further used to screen for small molecules that induce cell growth phenotypic changes affecting cell wall. Identification of such small molecules is beneficial due to their posttranslational mechanism of action that can be controlled in a temporal and spatial manner. Chemical genomics has the ability to overcome issues of genetic redundancy and lethality, which can hinder traditional genetic methods. The identification of small molecules in these screens will provide useful information on plant cell wall biology and overall plant growth.

The structure and biochemistry of charophycean cell walls: I. Pectins of Penium margaritaceum.

Plant cell walls are essential for proper growth, development, and interaction with the environment. It is generally accepted that land plants arose from aquatic ancestors which are sister groups to the charophycean algae (i.e., Streptophyta), and study of wall evolution during this transition promises insight into structure-function relationships of wall components. In this paper, we explore wall evolutionary history by studying the incorporation of pectin polymers into cell walls of the model organism Penium margaritaceum, a simple single-cell desmid. This organism produces only a primary wall consisting of three fibrillar or fibrous layers, with the outermost stratum terminating in distinct, calcified projections. Extraction of isolated cell walls with trans-1,2-diaminocyclohexane-N,N,N',N'-tetraacetic acid yielded a homogalacturonan (HGA) that was partially methyl esterified and equivalent to that found in land plants. Other pectins common to land plants were not detected, although selected components of some of these polymers were present. Labeling with specific monoclonal antibodies raised against higher-plant HGA epitopes (e.g., JIM5, JIM7, LM7, 2F4, and PAM1) demonstrated that the wall complex and outer layer projections were composed of the HGA which was significantly calcium complexed. JIM5 and JIM7 labeling suggested that highly methyl esterified HGA was secreted into the isthmus zone of dividing cells, the site of active wall secretion. As the HGA was displaced to more polar regions, de-esterification in a non-blockwise fashion occurred. This, in turn, allowed for calcium binding and the formation of the rigid outer wall layer. The patterning of HGA deposition provides interesting insights into the complex process of pectin involvement in the development of the plant cell wall.

Development of the cell wall in Tetraselmis: role of the Golgi apparatus and extracellular wall assembly.

The green algal flagellate, Tetraselmis, is a key transition organism in the phylogeny of green algae. It has been proposed that the cell wall of Tetraselmis arose evolutionarily from the fusion of scales and that this event secondarily caused the alteration of some cytoplasmic processes such as mitotic and cytokinetic mechanisms. Ultrastructural and developmental studies of the cell wall were performed with several strains of Tetraselmis. Two major wall types are reported. The wall of type 1 cells consists of a thick inner region covered by a layer of regularly repeating subunits of 26 nm, comparable to the subunits found in the median W2-W6 layer of Chlamydomonas. The more elaborate type 2 cell wall consists of a thick median wall layer, homologous to the type 1 inner wall, with additional inner and outer strata of hairs, grains and scales. Development of the cell wall begins in the endomembrane system, particularly the Golgi apparatus, where fibrillar tufts and electron-dense droplets are synthesized, modified and transported to the outside. Here, the tufts and droplets are displaced around the protoplast and assemble in several steps to yield the intact wall. Edge-growth assembly of the wall occurs here synchronously with cytoplasmic developments to yield the characteristic anterior flagellar pit. Models explaining various aspects of this development are discussed. When released from the cell, the wall subunits are not completely comparable to stellate scales, but appear to correspond to developmental stages of scales in green flagellates possessing body scales.

The comparative aspects of cell wall chemistry in the green algae (Chlorophyta).

The origin of a cell wall was an event of fundamental importance in the evolution of plants. In the green algae, cell walls apparently had independent origins in at least three lines of evolution. In this paper, the components of the cell wall were determined and compared in four filamentous green algae representing the charophycean, chlorophycean and ulvacean evoluationary lines. The walls of all four have hydroxyproline-containing proteins which separate into five or six bands upon SDS gel electrophoresis. Variation does exist, with the charophyte possessing fast moving electrophoretic bands and high hydroxyproline content, the chlorophytes having intermediate movement of bands and lower hydroxyproline content, and the ulvaecean representative possessing slow moving bands and a very low, if not questionable, hydroxyproline and saccharide content. Qualitative and quantitative estimates of wall proteins and sugars have been determined and compared. A hypothetical scheme of cell wall evolution based on these data, those of previous analyses, and recent phylogenetic schemes is presented. Although sound conclusions cannot be made until more information is available, the scheme might help to emphasize the areas most in need of additional research.

Mechanotransduction molecules in the plant gravisensory response: amyloplast/statolith membranes contain a beta 1 integrin-like protein.

It has been hypothesized that the sedimentation of amyloplasts within root cap cells is the primary event in the plant gravisensory-signal transduction cascade. Statolith sedimentation, with its ability to generate weighty mechanical signals, is a legitimate means for organisms to discriminate the direction of the gravity vector. However, it has been demonstrated that starchless mutants with reduced statolith densities maintain some ability to sense gravity, calling into question the statolith sedimentation hypothesis. Here we report on the presence of a beta 1 integrin-like protein localized inside amyloplasts of tobacco NT-1 suspension culture, callus cells, and whole-root caps. Two different antibodies to the beta 1 integrin, one to the cytoplasmic domain and one to the extracellular domain, localize in the vicinity of the starch grains within amyloplasts of NT-1. Biochemical data reveals a 110-kDa protein immunoprecipitated from membrane fractions of NT-1 suspension culture indicating size homology to known beta 1 integrin in animals. This study provides the first direct evidence for the possibility of integrin-mediated signal transduction in the perception of gravity by higher plants. An integrin-mediated pathway, initiated by starch grain sedimentation within the amyloplast, may provide the signal amplification necessary to explain the gravitropic response in starch-depleted cultivars.