Rice OVATE family protein 6 regulates leaf angle by modulating secondary cell wall biosynthesis

Plant Molecular Biology - Secondary cell wall not only provides rigidity and mechanical resistance to plants, but also has a large impact on plant growth and adaptation to environments....     

Auxin-induced cell elongation and cell wall changes

It has been well known that auxin induces cell elongation through its effect on modifications of the cell wall. The present review will discuss cell wall modifications, physical and biochemical, as the background of the former, based on the experimental results from our laboratory and from others, with the historical background. Discussions will particularly put stress on the auxin effect on the cell wall in terms of the following studies, namely, (1) measurements of the mechanical property of the cell wall, and (2) biochemical studies on the polysaccharide molecules of the cell wall. This article is dedicated to Professor Anton N.J. Heyn for his 85th birthday.     

Candida albicans protein kinase CaHsl1p regulates cell elongation and virulence

Saccharomyces cerevisiae Hsl1p is a Ser/Thr protein kinase that regulates cell morphology. We identified Candida albicans CaHSL1 and analysed its function in C. albicans. Cells lacking CaHsl1p exhibited filamentous growth under yeast growth conditions with the filaments elongating more quickly than did those of the wild type under hyphal growth conditions, suggesting that it plays a role in the suppression of cell elongation. Green fluorescent protein-tagged CaHsl1p colocalized with a septin complex to the bud neck during yeast growth or to a potent septation site during hyphal growth, as expected from the localization in S. cerevisiae. However, the localization of the septin complex did not change in DeltaCahsl1, suggesting that CaHsl1p does not participate in septin organization. CaHsl1p was expressed in a cell cycle-dependent manner and, except for the G1 phase, phosphorylated throughout the cell cycle. In DeltaCahsl1 cells, the phosphorylation of a possible CaHsl1p target CaSwe1p decreased, while that of CaCdc28p at tyrosine18 increased. Either an extra copy of the tyrosine18-mutated CaCdc28p or deletion of CaSWE1 suppressed the cell elongation phenotype caused by CaHSL1 deletion. Furthermore, DeltaCahsl1 exhibited reduced virulence in the mouse systemic candidiasis model. Thus, the CaHsl1p-CaSwe1p-CaCdc28p pathway appears important in the cell elongation of both the yeast and hyphal forms and to the virulence of C. albicans.     

Schizosaccharomyces pombe rho2p GTPase regulates cell wall alpha-glucan biosynthesis through the protein kinase pck2p

Schizosaccharomyces pombe rho1(+) and rho2(+) genes are involved in the control of cell morphogenesis, cell integrity, and polarization of the actin cytoskeleton. Although both GTPases interact with each of the two S. pombe protein kinase C homologues, Pck1p and Pck2p, their functions are distinct from each other. It is known that Rho1p regulates (1,3)beta-D-glucan synthesis both directly and through Pck2p. In this paper, we have investigated Rho2p signaling and show that pck2 delta and rho2 delta strains display similar defects with regard to cell wall integrity, indicating that they might be in the same signaling pathway. We also show that Rho2 GTPase regulates the synthesis of alpha-D-glucan, the other main structural polymer of the S. pombe cell wall, primarily through Pck2p. Although overexpression of rho2(+) in wild-type or pck1 delta cells is lethal and causes morphological alterations, actin depolarization, and an increase in alpha-D-glucan biosynthesis, all of these effects are suppressed in a pck2 delta strain. In addition, genetic interactions suggest that Rho2p and Pck2p are important for the regulation of Mok1p, the major (1-3)alpha-D-glucan synthase. Thus, a rho2 delta mutation, like pck2 delta, is synthetically lethal with mok1-664, and the mutant partially fails to localize Mok1p to the growing areas. Moreover, overexpression of mok1(+) in rho2 delta cells causes a lethal phenotype that is completely different from that of mok1(+) overexpression in wild-type cells, and the increase in alpha-glucan is considerably lower. Taken together, all of these results indicate the presence of a signaling pathway regulating alpha-glucan biosynthesis in which the Rho2p GTPase activates Pck2p, and this kinase in turn controls Mok1p.     

Polysaccharidases and the control of cell wall elongation

Summary Avena coleoptile sections were treated with a fraction of a fungal filtrate containing a potent cellulase. Elongation rate was not affected although turgor pressure remained constant and wall extensibility was increased. These data show that the simple weakening of cell walls is not sufficient to promote growth and suggest that endogenous polysaccharidases are not the means by which the growth rate of the coleoptile is regulated.This work was supported in part by a predoctoral and a postdoctoral fellowship from NSF     

A plasma membrane-bound putative endo-1,4-beta-D-glucanase is required for normal wall assembly and cell elongation in Arabidopsis.

Endo-1,4-beta-D-glucanases (EGases) form a large family of hydrolytic enzymes in prokaryotes and eukaryotes. In higher plants, potential substrates in vivo are xyloglucan and non-crystalline cellulose in the cell wall. Gene expression patterns suggest a role for EGases in various developmental processes such as leaf abscission, fruit ripening and cell expansion. Using Arabidopsis thaliana genetics, we demonstrate the requirement of a specialized member of the EGase family for the correct assembly of the walls of elongating cells. KORRIGAN (KOR) is identified by an extreme dwarf mutant with pronounced architectural alterations in the primary cell wall. The KOR gene was isolated and encodes a membrane-anchored member of the EGase family, which is highly conserved between mono- and dicotyledonous plants. KOR is located primarily in the plasma membrane and presumably acts at the plasma membrane-cell wall interface. KOR mRNA was found in all organs examined, and in the developing dark-grown hypocotyl, mRNA levels were correlated with rapid cell elongation. Among plant growth factors involved in the control of hypocotyl elongation (auxin, gibberellins and ethylene) none significantly influenced KOR-mRNA levels. However, reduced KOR-mRNA levels were observed in det2, a mutant deficient for brassinosteroids. Although the in vivo substrate remains to be determined, the mutant phenotype is consistent with a central role for KOR in the assembly of the cellulose-hemicellulose network in the expanding cell wall.     

Bridging cell wall biosynthesis and bacterial morphogenesis

The bacterial cell wall is a complex three-dimensional structure that protects the cell from environmental stress and ensures its shape. The biosynthesis of its main component, the peptidoglycan, involves the coordination of activities of proteins present in the cytoplasm, the membrane, and the periplasm, some of which also interact with the bacterial cytoskeleton. The sheer complexity of the cell wall elongation process, which is the main focus of this review, has created a significant challenge for the study of the macromolecular interactions that regulate peptidoglycan biosynthesis. The availability of new structural and biochemical data on a number of components of peptidoglycan assembly machineries, including a complex between MreB and RodZ as well as structures of penicillin-binding proteins (PBPs) from a number of pathogenic species, now provide novel insight into the underpinnings of an intricate molecular machinery.     

Codon usage and protein length-dependent feedback from translation elongation regulates translation initiation and elongation speed

Essential cellular functions require efficient production of many large proteins but synthesis of large proteins encounters many obstacles in cells. Translational control is mostly known to be regulated at the initiation step. Whether translation elongation process can feedback to regulate initiation efficiency is unclear. Codon usage bias, a universal feature of all genomes, plays an important role in determining gene expression levels. Here, we discovered that there is a conserved but codon usage-dependent genome-wide negative correlation between protein abundance and CDS length. The codon usage effects on protein expression and ribosome flux on mRNAs are influenced by CDS length; optimal codon usage preferentially promotes production of large proteins. Translation of mRNAs with long CDS and non-optimal codon usage preferentially induces phosphorylation of initiation factor eIF2α, which inhibits translation initiation efficiency. Deletion of the eIF2α kinase CPC-3 (GCN2 homolog) in Neurospora preferentially up-regulates large proteins encoded by non-optimal codons. Surprisingly, CPC-3 also inhibits translation elongation rate in a codon usage and CDS length-dependent manner, resulting in slow elongation rates for long CDS mRNAs. Together, these results revealed a codon usage and CDS length-dependent feedback mechanism from translation elongation to regulate both translation initiation and elongation kinetics.     

Stress relaxation property of the cell wall and auxin-induced cell elongation

A stress-relaxation method has been developed to measure the mechanical property of the plant cell wall, as a physically defined terms. In the method, the stress relaxation property of the cell wall is simulated with a Maxwell viscoelastic model whose character is represented by four parameters; the minimum relaxation time, To, the relaxation rate, b, the maximum relaxation time, Tm and the residual stress, c. Thus, the mechanical property of the cell wall is represented by the four parameters. Physical and physiological meanings of the parameters are discussed. Auxin effects on the parameters were also studied. The cell elongation is simply thought to be extension of the cell wall under a force. The extension of the cell wall can be simulated by the mechanical property of the cell wall. However, the calculated extension was found to be incomparable to the real cell growth, indicating that there has to be other factors limiting the rate of cell growth. Major factors governing cell growth are discussed to be the cell wall mechanical property, the osmotic potential and water movement in the apoplast. A possibility to predict cell expansion with the three factors was discussed and a novel equation representing cell growth was obtained: $$1/R = 1/R_w + 1/R_p $$ whereR is the rate of cell elongation,R _w is the rate of cell wall extension due to the osmotic pressure andR _p is the rate of cell elongation determined by water conductivity.     

A novel screening method for cell wall mutants in Aspergillus niger identifies UDP-galactopyranose mutase as an important protein in fungal cell wall biosynthesis

To identify cell wall biosynthetic genes in filamentous fungi and thus potential targets for the discovery of new antifungals, we developed a novel screening method for cell wall mutants. It is based on our earlier observation that the Aspergillus niger agsA gene, which encodes a putative alpha-glucan synthase, is strongly induced in response to cell wall stress. By placing the agsA promoter region in front of a selectable marker, the acetamidase (amdS) gene of A. nidulans, we reasoned that cell wall mutants with a constitutively active cell wall stress response pathway could be identified by selecting mutants for growth on acetamide as the sole nitrogen source. For the genetic screen, a strain was constructed that contained two reporter genes controlled by the same promoter: the metabolic reporter gene PagsA-amdS and PagsA-H2B-GFP, which encodes a GFP-tagged nuclear protein. The primary screen yielded 161 mutants that were subjected to various cell wall-related secondary screens. Four calcofluor white-hypersensitive, osmotic-remediable thermosensitive mutants were selected for complementation analysis. Three mutants were complemented by the same gene, which encoded a protein with high sequence identity with eukaryotic UDP-galactopyranose mutases (UgmA). Our results indicate that galactofuranose formation is important for fungal cell wall biosynthesis and represents an attractive target for the development of antifungals.     

OsDOG, a gibberellin-induced A20/AN1 zinc-finger protein, negatively regulates gibberellin-mediated cell elongation in rice

The A20/AN1 zinc-finger proteins (ZFPs) play pivotal roles in animal immune responses and plant stress responses. From previous gibberellin (GA) microarray data and A20/AN1 ZFP family member association, we chose Oryza sativa dwarf rice with overexpression of gibberellin-induced gene (OsDOG) to examine its function in the GA pathway. OsDOG was induced by gibberellic acid (GA₃) and repressed by the GA-synthesis inhibitor paclobutrazol. Different transgenic lines with constitutive expression of OsDOG showed dwarf phenotypes due to deficiency of cell elongation. Additional GA₁ and real-time PCR quantitative assay analyses confirmed that the decrease of GA₁ in the overexpression lines resulted from reduced expression of GA3ox2 and enhanced expression of GA2ox1 and GA2ox3. Adding exogenous GA rescued the constitutive expression phenotypes of the transgenic lines. OsDOG has a novel function in regulating GA homeostasis and in negative maintenance of plant cell elongation in rice.     

Hijacking of a substrate-binding protein scaffold for use in mycobacterial cell wall biosynthesis

The waxy cell wall is crucial to the survival of mycobacteria within the infected host. The cell wall is a complex structure rich in unusual molecules that includes two related lipoglycans, the phosphatidylinositol mannosides (PIMs) and lipoarabinomannans (LAMs). Many proteins implicated in the PIM/LAM biosynthetic pathway, while attractive therapeutic targets, are poorly defined. The 2.4A resolution crystal structure of an essential lipoprotein, LpqW, implicated in LAM biosynthesis is reported here. LpqW adopts a scaffold reminiscent of the distantly related, promiscuous substrate-binding proteins of the ATP-binding cassette import system. Nevertheless, the unique closed conformation of LpqW suggests that mycobacteria and other closely related pathogens have hijacked this scaffold for use in key processes of cell wall biosynthesis. In silico docking provided a plausible model in which the candidate PIM ligand binds within a marked electronegative region located on the surface of LpqW. We suggest that LpqW represents an archetypal lipoprotein that channels intermediates from a pathway for mature PIM production into a pathway for LAM biosynthesis, thus controlling the relative abundance of these two important components of the cell wall.     

Rice cell wall polysaccharides: Structure and biosynthesis

Rice is the most widely consumed staple food, and is cultivated worldwide to satisfy our daily caloric needs. Thus, extensive efforts on rice breeding and biotechnology have substantially focused on the development of elite cultivars with high yields and better grain quality, as well as enhanced resistance against biotic and abiotic stresses. Recently, it has been observed that rice is more than a just grain-producing crop. Carbon-rich materials of the rice cell wall polysaccharides from post-harvest wastes, including the straw and husk, have been converted into bioethanol and other invaluable, renewable materials. In order to maximize the utilization of cell wall-derived resources, it is imperative to understand cell wall chemistry and molecular mechanism underlying cell wall biosynthesis in rice. In the last decade, several approaches, including mutational genetics and the functional characterization of candidate genes, have been successful in isolating some of cell wall biosynthetic genes in rice, marking the first step forward in obtaining a complete understanding of rice cell wall biosynthesis, although the exact biochemical functions have not been conclusive. In this paper, we focus on integrating old and new information to provide an updated perspective in the cell wall formation of rice, highlighting the chemical structures and biosynthesis of rice cell wall polysaccharides.