title: Human substance P receptor lacking the C-terminal domain remains competent to desensitize and internalize.
abstract: Substance P receptor (SPR) and its naturally occurring splice-variant, lacking the C-terminal tail, are found in brain and spinal cord. Whether C-terminally truncated SPR desensitizes like full-length SPR is controversial. We used a multivaried approach to determine whether human SPR (hSPR) and a C-terminally truncated mutant, hSPRDelta325, differ in their desensitization and internalization. In HEK-293 cells expressing either hSPRDelta325 or hSPR, SP-induced desensitization of the two receptors was similar when measured by inositol triphosphate accumulation or by transient translocation of coexpressed PKCbetaII-GFP to the plasma membrane. Moreover, translocation of beta-arrestin 1 or 2-GFP (betaarr1-GFP or betaarr2-GFP) to the plasma membrane, and receptor internalization were also similar. However, hSPR and hSPRDelta325 differ in their phosphorylation and in their ability to form beta-arrestin-containing endocytic vesicles. Unlike hSPR, hSPRDelta325 is not phosphorylated to a detectable level in intact HEK293 cells, and whereas hSPR forms vesicles containing either betaarr1-GFP or betaarr2-GFP, hSPRDelta325 does not form any vesicles with betaarr1-GFP, and forms fewer vesicles with betaarr2-GFP. We conclude that truncated hSPR undergoes agonist-dependent desensitization and internalization without detectable receptor phosphorylation.
authors.affiliation: Departments of Anesthesiology and Cell Biology, The Howard Hughes Medical Institute, Duke University Medical Center, Durham, NC 27710, USA.
authors.forename: Mark D
authors.initials: MD
authors.lastname: Richardson
authors.forename: Anastasia M
authors.initials: AM
authors.lastname: Balius
authors.forename: Keisuke
authors.initials: K
authors.lastname: Yamaguchi
authors.forename: Emily R
authors.initials: ER
authors.lastname: Freilich
authors.forename: Larry S
authors.initials: LS
authors.lastname: Barak
authors.forename: Madan M
authors.initials: MM
authors.lastname: Kwatra
mesh_terms: D000595:Amino Acid Sequence; D019390:Arrestins / Q000235:genetics; D002460:Cell Line; D049452:Green Fluorescent Proteins; D006801:Humans; D007527:Isoenzymes / Q000235:genetics; D007668:Kidney / Q000166:cytology / Q000378:metabolism; D008164:Luminescent Proteins / Q000235:genetics; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D010766:Phosphorylation; D011485:Protein Binding / Q000502:physiology; D020033:Protein Isoforms / Q000235:genetics; D011493:Protein Kinase C / Q000235:genetics; D064546:Protein Kinase C beta; D017434:Protein Structure, Tertiary / Q000502:physiology; D021381:Protein Transport / Q000502:physiology; D018040:Receptors, Neurokinin-1 / Q000187:drug effects / Q000235:genetics* / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D017384:Sequence Deletion; D013373:Substance P / Q000494:pharmacology; D014162:Transfection; D000071559:beta-Arrestin 1; D000071557:beta-Arrestins
chemical_list: C000605253:ARRB1 protein, human; D019390:Arrestins; D007527:Isoenzymes; D008164:Luminescent Proteins; D020033:Protein Isoforms; D018040:Receptors, Neurokinin-1; D011993:Recombinant Fusion Proteins; D000071559:beta-Arrestin 1; D000071557:beta-Arrestins; D049452:Green Fluorescent Proteins; D013373:Substance P; D011493:Protein Kinase C; D064546:Protein Kinase C beta