Conformational Dynamics Underlie the Activity of the Auxin-binding Protein, Nt-abp1*

The auxin-binding protein 1 (ABP1) has been proposed to be involved in the perception of the phytohormone at the plasma membrane. Site-directed mutagenesis was performed on highly conserved residues at the C terminus of ABP1 to investigate their relative importance in protein folding and activation of a functional response at the plasma membrane. Detailed analysis of the dynamic interaction of the wild-type ABP1 and mutated proteins with three distinct monoclonal antibodies recognizing conformation-dependent epitopes was performed by surface plasmon resonance. The influence of auxin on these interactions was also investigated. The Cys177 as well as Asp175 and Glu176 were identified as critical residues for ABP1 folding and action at the plasma membrane. On the contrary, the C-terminal KDEL sequence was demonstrated not to be essential for auxin binding, interaction with the plasma membrane, or activation of the transduction cascade although it does appear to be involved in the stability of ABP1. Taken together, the results confirmed that ABP1 conformational change is the critical step for initiating the signal from the plasma membrane.

[1]  J. Grosclaude,et al.  Real-time Kinetic Measurements of the Interactions between Lactogenic Hormones and Prolactin-Receptor Extracellular Domains from Several Species Support the Model of Hormone-induced Transient Receptor Dimerization* , 1996, The Journal of Biological Chemistry.

[2]  J. Nemhauser,et al.  ETTIN patterns the Arabidopsis floral meristem and reproductive organs. , 1997, Development.

[3]  T. Arakawa,et al.  Dimerization of the extracellular domain of the erythropoietin (EPO) receptor by EPO: one high-affinity and one low-affinity interaction. , 1996, Biochemistry.

[4]  H. Pelham,et al.  Immunological evidence that plants use both HDEL and KDEL for targeting proteins to the endoplasmic reticulum. , 1992, Journal of cell science.

[5]  U. Dohrmann,et al.  Characterization of naphthaleneacetic Acid binding to receptor sites on cellular membranes of maize coleoptile tissue. , 1977, Plant physiology.

[6]  Klaus Palme,et al.  AtPIN2 defines a locus of Arabidopsis for root gravitropism control , 1998, The EMBO journal.

[7]  S. Loefas,et al.  Immobilization of proteins to a carboxymethyldextran-modified gold surface for biospecific interaction analysis in surface plasmon resonance sensors. , 1991, Analytical biochemistry.

[8]  M. Savka,et al.  Auxin-dependent cell expansion mediated by overexpressed auxin-binding protein 1. , 1998, Science.

[9]  M. Blatt,et al.  K+ channels of stomatal guard cells: bimodal control of the K+ inward-rectifier evoked by auxin. , 1994, The Plant journal : for cell and molecular biology.

[10]  M. Goldsmith.  Cellular signaling: new insights into the action of the plant growth hormone auxin. , 1993, Proceedings of the National Academy of Sciences of the United States of America.

[11]  J. Chory,et al.  Regulation of Auxin Response by the Protein Kinase PINOID , 2000, Cell.

[12]  G. Ephritikhine,et al.  Auxin effect on the transmembrane potential difference of wild-type and mutant tobacco protoplasts exhibiting a differential sensitiity to auxin. , 1987, Plant physiology.

[13]  D. Söll,et al.  The RCN1-encoded A subunit of protein phosphatase 2A increases phosphatase activity in vivo. , 1999, The Plant journal : for cell and molecular biology.

[14]  G. Fink,et al.  EIR1, a root-specific protein involved in auxin transport, is required for gravitropism in Arabidopsis thaliana. , 1998, Genes & development.

[15]  J. Grosclaude,et al.  A Novel Immunological Approach Establishes That the Auxin-binding Protein, Nt-abp1, Is an Element Involved in Auxin Signaling at the Plasma Membrane* , 1999, The Journal of Biological Chemistry.

[16]  P. Masson,et al.  The arabidopsis thaliana AGRAVITROPIC 1 gene encodes a component of the polar-auxin-transport efflux carrier. , 1998, Proceedings of the National Academy of Sciences of the United States of America.

[17]  M. Fricker,et al.  Modulation of K+ channels in Vicia stomatal guard cells by peptide homologs to the auxin-binding protein C terminus. , 1993, Proceedings of the National Academy of Sciences of the United States of America.

[18]  W. Stiekema,et al.  Improving scFv antibody expression levels in the plant cytosol 1 , 1997, FEBS letters.

[19]  Alan M. Jones,et al.  Auxin-Binding Proteins , 1994 .

[20]  K. Palme,et al.  Molecular analysis of an auxin binding protein gene located on chromosome 4 of Arabidopsis. , 1992, The Plant cell.

[21]  C. Gehring,et al.  Auxin-binding-protein antibodies and peptides influence stomatal opening and alter cytoplasmic pH , 1998, Planta.

[22]  M. Futai,et al.  Purification and properties of an auxin-binding protein from maize shoot membranes. , 1986, Journal of biochemistry.

[23]  M. A. Venis,et al.  Auxin Receptors and Auxin Binding Proteins , 1995 .

[24]  F. B. Pickett,et al.  Mutations in the AXR3 gene of Arabidopsis result in altered auxin response including ectopic expression from the SAUR-AC1 promoter. , 1996, The Plant journal : for cell and molecular biology.

[25]  J. Vandekerckhove,et al.  Molecular cloning and structural analysis of a gene from Zea mays (L.) coding for a putative receptor for the plant hormone auxin. , 1989, The EMBO journal.

[26]  C. Hardtke,et al.  The Arabidopsis gene MONOPTEROS encodes a transcription factor mediating embryo axis formation and vascular development , 1998, The EMBO journal.

[27]  M. Estelle,et al.  Function of the ubiquitin-proteasome pathway in auxin response. , 2000, Trends in biochemical sciences.

[28]  C. Perrot-Rechenmann,et al.  The auxin‐binding protein Nt‐ERabp1 alone activates an auxin‐like transduction pathway , 1999, FEBS letters.

[29]  M Löbler,et al.  Auxin-binding protein from coleoptile membranes of corn (Zea mays L.). I. Purification by immunological methods and characterization. , 1985, The Journal of biological chemistry.

[30]  Michael A. Venis,et al.  OPINIONAuxin binding protein 1 is a red herring? Oh no it isn't! , 1995 .

[31]  M. Venis.  Solubilisation and partial purification of auxin-binding sites of corn membranes , 1977, Nature.

[32]  K. Feldmann,et al.  Arabidopsis AUX1 Gene: A Permease-Like Regulator of Root Gravitropism , 1996, Science.

[33]  H. Pelham,et al.  The retention signal for soluble proteins of the endoplasmic reticulum. , 1990, Trends in biochemical sciences.

[34]  W. Diekmann,et al.  Auxins induce clustering of the auxin-binding protein at the surface of maize coleoptile protoplasts. , 1995, Proceedings of the National Academy of Sciences of the United States of America.

[35]  R. Hertel,et al.  OPINION: Auxin binding protein 1 is a red herring , 1995 .