Elucidation of the Signal Transduction Pathways Activated by the Plant Natriuretic Peptide AtPNP-A

Handle URI:
http://hdl.handle.net/10754/335803
Title:
Elucidation of the Signal Transduction Pathways Activated by the Plant Natriuretic Peptide AtPNP-A
Authors:
Turek, Ilona ( 0000-0001-6457-7456 )
Abstract:
Plant natriuretic peptides (PNPs) comprise a novel class of hormones that share some sequence similarity in the active site with their animal analogues that function as regulators of salt and water balance. A PNP present in Arabidopsis thaliana (AtPNP-A) has been assigned a role in abiotic and biotic stress responses, and the recombinant protein has been demonstrated to elicit cyclic guanosine monophosphate (cGMP)-dependent stomatal guard cell opening, regulate ion movements, and induce osmoticum-dependent water uptake. Although the importance of the hormone in maintaining ion and fluid homeostasis has been established, key components of the AtPNP-A-dependent signal transduction pathway remain unknown. Since identification of the binding partners of AtPNP-A, including its receptor(s), is fundamental to understanding the mode of its action at the molecular level, comprehensive protein-protein interaction studies, involving yeast two-hybrid screening, affinity-based assays, protein cross-linking and co-immunoprecipitation followed by mass spectrometric (MS) analyses have been performed. Several candidate binding partners of AtPNP-A identified with at least two independent methods were subsequently expressed as recombinant proteins, purified, and the specificity of their interactions with the recombinant AtPNP-A was verified using surface plasmon resonance. Several specific binary interactants of AtPNP-A were subjected to functional assays aimed at unraveling the consequences of the interactions in planta. These experiments have revealed that reactive oxygen species (ROS) are novel secondary messengers involved in the transduction of AtPNP-A signal in suspension-cultured cells of A. thaliana (Col-0). Further insight into the AtPNP-A dependent signalling events occurring in suspension-cultured cells in ROS-dependent or ROS-independent manner have been obtained from the large-scale proteomics study employing tandem mass tag (TMT) labelling followed by MS analysis to identify and relatively quantify proteins that are differentially expressed upon the treatment with nano- and picomolar concentrations of the biologically active AtPNP-A peptide at different time-points post-treatment. Characterization of both the AtPNP-A interactome and AtPNP-A dependent proteome afforded novel insights into the signal transduction pathways altered by PNPs and shed new light on the mechanisms by which these candidate interactants operate. Taken together, indications are that PNP dependent mechanisms can be harnessed for possible biotechnological applications.
Advisors:
Gehring, Christoph A ( 0000-0003-4355-4591 )
Committee Member:
Ravasi, Timothy ( 0000-0002-9950-465X ) ; Xiong, Liming ( 0000-0001-8099-0806 ) ; Elia, Giuliano ( 0000-0001-8216-9560 ) ; Lai, Zhiping ( 0000-0001-9555-6009 )
KAUST Department:
Physical Sciences and Engineering (PSE) Division
Program:
Chemical and Biological Engineering
Issue Date:
Nov-2014
Type:
Dissertation
Appears in Collections:
Dissertations; Physical Sciences and Engineering (PSE) Division; Chemical and Biological Engineering Program

Full metadata record

DC FieldValue Language
dc.contributor.advisorGehring, Christoph Aen
dc.contributor.authorTurek, Ilonaen
dc.date.accessioned2014-11-19T12:13:32Z-
dc.date.available2014-11-19T12:13:32Z-
dc.date.issued2014-11en
dc.identifier.urihttp://hdl.handle.net/10754/335803en
dc.description.abstractPlant natriuretic peptides (PNPs) comprise a novel class of hormones that share some sequence similarity in the active site with their animal analogues that function as regulators of salt and water balance. A PNP present in Arabidopsis thaliana (AtPNP-A) has been assigned a role in abiotic and biotic stress responses, and the recombinant protein has been demonstrated to elicit cyclic guanosine monophosphate (cGMP)-dependent stomatal guard cell opening, regulate ion movements, and induce osmoticum-dependent water uptake. Although the importance of the hormone in maintaining ion and fluid homeostasis has been established, key components of the AtPNP-A-dependent signal transduction pathway remain unknown. Since identification of the binding partners of AtPNP-A, including its receptor(s), is fundamental to understanding the mode of its action at the molecular level, comprehensive protein-protein interaction studies, involving yeast two-hybrid screening, affinity-based assays, protein cross-linking and co-immunoprecipitation followed by mass spectrometric (MS) analyses have been performed. Several candidate binding partners of AtPNP-A identified with at least two independent methods were subsequently expressed as recombinant proteins, purified, and the specificity of their interactions with the recombinant AtPNP-A was verified using surface plasmon resonance. Several specific binary interactants of AtPNP-A were subjected to functional assays aimed at unraveling the consequences of the interactions in planta. These experiments have revealed that reactive oxygen species (ROS) are novel secondary messengers involved in the transduction of AtPNP-A signal in suspension-cultured cells of A. thaliana (Col-0). Further insight into the AtPNP-A dependent signalling events occurring in suspension-cultured cells in ROS-dependent or ROS-independent manner have been obtained from the large-scale proteomics study employing tandem mass tag (TMT) labelling followed by MS analysis to identify and relatively quantify proteins that are differentially expressed upon the treatment with nano- and picomolar concentrations of the biologically active AtPNP-A peptide at different time-points post-treatment. Characterization of both the AtPNP-A interactome and AtPNP-A dependent proteome afforded novel insights into the signal transduction pathways altered by PNPs and shed new light on the mechanisms by which these candidate interactants operate. Taken together, indications are that PNP dependent mechanisms can be harnessed for possible biotechnological applications.en
dc.language.isoenen
dc.subjectplant natriuretic peptidesen
dc.subjectplant stress signalingen
dc.subjectProtein-protein interactionsen
dc.subjectsecondary messengersen
dc.subjectcyclic guanosine monophosphateen
dc.subjectArabidopsis Thalianaen
dc.titleElucidation of the Signal Transduction Pathways Activated by the Plant Natriuretic Peptide AtPNP-Aen
dc.typeDissertationen
dc.contributor.departmentPhysical Sciences and Engineering (PSE) Divisionen
thesis.degree.grantorKing Abdullah University of Science and Technologyen_GB
dc.contributor.committeememberRavasi, Timothyen
dc.contributor.committeememberXiong, Limingen
dc.contributor.committeememberElia, Giulianoen
dc.contributor.committeememberLai, Zhipingen
thesis.degree.disciplineChemical and Biological Engineeringen
thesis.degree.nameDoctor of Philosophyen
dc.person.id101752en
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