title: Interaction of receptor-activity-modifying protein1 with tubulin.
abstract: Receptor-activity-modifying protein (RAMP) 1 is an accessory protein of the G protein-coupled calcitonin receptor-like receptor (CLR). The CLR/RAMP1 heterodimer defines a receptor for the potent vasodilatory calcitonin gene-related peptide. A wider tissue distribution of RAMP1, as compared to that of the CLR, is consistent with additional biological functions. Here, glutathione S-transferase (GST) pull-down, coimmunoprecipitation and yeast two-hybrid experiments identified beta-tubulin as a novel RAMP1-interacting protein. GST pull-down experiments indicated interactions between the N- and C-terminal domains of RAMP1 and beta-tubulin. Yeast two-hybrid experiments confirmed the interaction between the N-terminal region of RAMP1 and beta-tubulin. Interestingly, alpha-tubulin was co-extracted with beta-tubulin in pull-down experiments and immunoprecipitation of RAMP1 coprecipitated alpha- and beta-tubulin. Confocal microscopy indicated colocalization of RAMP1 and tubulin predominantly in axon-like processes of neuronal differentiated human SH-SY5Y neuroblastoma cells. In conclusion, the findings point to biological roles of RAMP1 beyond its established interaction with G protein-coupled receptors.
authors.affiliation: Research Laboratory for Calcium Metabolism, Orthopedic University Hospital Balgrist, University of Zurich, 8008 Zurich, Switzerland.
authors.forename: Thomas H
authors.initials: TH
authors.lastname: Kunz
authors.forename: Sarah
authors.initials: S
authors.lastname: Mueller-Steiner
authors.forename: Kerstin
authors.initials: K
authors.lastname: Schwerdtfeger
authors.forename: Peter
authors.initials: P
authors.lastname: Kleinert
authors.forename: Heinz
authors.initials: H
authors.lastname: Troxler
authors.forename: Jens M
authors.initials: JM
authors.lastname: Kelm
authors.forename: Lars M
authors.initials: LM
authors.lastname: Ittner
authors.forename: Jan A
authors.initials: JA
authors.lastname: Fischer
authors.forename: Walter
authors.initials: W
authors.lastname: Born
mesh_terms: D000957:Antigens, Viral, Tumor / Q000502:physiology; D001369:Axons / Q000378:metabolism; D002461:Cell Line, Transformed; D045744:Cell Line, Tumor; D002472:Cell Transformation, Viral; D003001:Cloning, Molecular; D004926:Escherichia coli / Q000235:genetics; D005982:Glutathione Transferase / Q000378:metabolism; D006801:Humans; D047908:Intracellular Signaling Peptides and Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D008565:Membrane Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D018613:Microscopy, Confocal; D009447:Neuroblastoma / Q000473:pathology; D009474:Neurons / Q000378:metabolism; D011233:Precipitin Tests; D017434:Protein Structure, Tertiary; D011506:Proteins / Q000378:metabolism*; D058262:Receptor Activity-Modifying Protein 1; D058261:Receptor Activity-Modifying Proteins; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D013539:Simian virus 40 / Q000502:physiology; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D014404:Tubulin / Q000737:chemistry / Q000378:metabolism*; D020798:Two-Hybrid System Techniques
chemical_list: D000957:Antigens, Viral, Tumor; D047908:Intracellular Signaling Peptides and Proteins; D008565:Membrane Proteins; D011506:Proteins; C545502:RAMP1 protein, human; D058262:Receptor Activity-Modifying Protein 1; D058261:Receptor Activity-Modifying Proteins; D011993:Recombinant Fusion Proteins; D014404:Tubulin; D005982:Glutathione Transferase