title: Tandem affinity purification of AtTERT reveals putative interaction partners of plant telomerase in vivo.
abstract: The life cycle of telomerase involves dynamic and complex interactions between proteins within multiple macromolecular networks. Elucidation of these associations is a key to understanding the regulation of telomerase under diverse physiological and pathological conditions from telomerase biogenesis, through telomere recruitment and elongation, to its non-canonical activities outside of telomeres. We used tandem affinity purification coupled to mass spectrometry to build an interactome of the telomerase catalytic subunit AtTERT, using Arabidopsis thaliana suspension cultures. We then examined interactions occurring at the AtTERT N-terminus, which is thought to fold into a discrete domain connected to the rest of the molecule via a flexible linker. Bioinformatic analyses revealed that interaction partners of AtTERT have a range of molecular functions, a subset of which is specific to the network around its N-terminus. A significant number of proteins co-purifying with the N-terminal constructs have been implicated in cell cycle and developmental processes, as would be expected of bona fide regulatory interactions and we have confirmed experimentally the direct nature of selected interactions. To examine AtTERT protein-protein interactions from another perspective, we also analysed AtTERT interdomain contacts to test potential dimerization of AtTERT. In total, our results provide an insight into the composition and architecture of the plant telomerase complex and this will aid in delineating molecular mechanisms of telomerase functions.
authors.affiliation: Institute of Biophysics, Academy of Sciences of the Czech Republic, v.v.i., Královopolská 135, CZ-61265, Brno, Czech Republic.|Central European Institute of Technology and Faculty of Science, Masaryk University, Kotlářská 2, CZ-61137, Brno, Czech Republic.|Swiss Institute for Experimental Cancer Research (ISREC), School of Life Sciences, École Polytechnique Fédérale de Lausanne (EPFL), 1015, Lausanne, Switzerland.
authors.forename: Jana
authors.initials: J
authors.lastname: Majerská
authors.affiliation: Central European Institute of Technology and Faculty of Science, Masaryk University, Kotlářská 2, CZ-61137, Brno, Czech Republic.|Swiss Institute for Experimental Cancer Research (ISREC), School of Life Sciences, École Polytechnique Fédérale de Lausanne (EPFL), 1015, Lausanne, Switzerland.
authors.forename: Petra Procházková
authors.initials: PP
authors.lastname: Schrumpfová
authors.affiliation: Institute of Biophysics, Academy of Sciences of the Czech Republic, v.v.i., Královopolská 135, CZ-61265, Brno, Czech Republic.
authors.forename: Ladislav
authors.initials: L
authors.lastname: Dokládal
authors.affiliation: Central European Institute of Technology and Faculty of Science, Masaryk University, Kotlářská 2, CZ-61137, Brno, Czech Republic.
authors.forename: Šárka
authors.initials: Š
authors.lastname: Schořová
authors.affiliation: Central European Institute of Technology and Faculty of Science, Masaryk University, Kotlářská 2, CZ-61137, Brno, Czech Republic.
authors.forename: Karel
authors.initials: K
authors.lastname: Stejskal
authors.affiliation: Central European Institute of Technology and Faculty of Science, Masaryk University, Kotlářská 2, CZ-61137, Brno, Czech Republic.
authors.forename: Michal
authors.initials: M
authors.lastname: Obořil
authors.affiliation: Institute of Experimental Biology, Academy of Sciences of the Czech Republic, v.v.i., Rozvojová 263, CZ-165 02, Prague, Czech Republic.
authors.forename: David
authors.initials: D
authors.lastname: Honys
authors.affiliation: Central European Institute of Technology and Faculty of Science, Masaryk University, Kotlářská 2, CZ-61137, Brno, Czech Republic.
authors.forename: Lucie
authors.initials: L
authors.lastname: Kozáková
authors.affiliation: Central European Institute of Technology and Faculty of Science, Masaryk University, Kotlářská 2, CZ-61137, Brno, Czech Republic.
authors.forename: Pavla Sováková
authors.initials: PS
authors.lastname: Polanská
authors.affiliation: Institute of Biophysics, Academy of Sciences of the Czech Republic, v.v.i., Královopolská 135, CZ-61265, Brno, Czech Republic. evin@ibp.cz.
authors.forename: Eva
authors.initials: E
authors.lastname: Sýkorová
mesh_terms: D017360:Arabidopsis / Q000201:enzymology* / Q000235:genetics; D029681:Arabidopsis Proteins / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism; D002467:Cell Nucleus / Q000201:enzymology; D002478:Cells, Cultured; D002846:Chromatography, Affinity; D015870:Gene Expression; D054730:Protein Interaction Domains and Motifs; D025941:Protein Interaction Mapping; D060066:Protein Interaction Maps; D055503:Protein Multimerization; D053719:Tandem Mass Spectrometry; D019098:Telomerase / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism
chemical_list: D029681:Arabidopsis Proteins; D019098:Telomerase
keywords: AtPOT1a; PURα1; Pontin; Reptin; TAP-MS; Telomerase