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Site-specific polyubiquitination differentially regulates parathyroid hormone receptor-initiated MAPK signaling and cell proliferation.

Publication ,  Journal Article
Zhang, Q; Xiao, K; Liu, H; Song, L; McGarvey, JC; Sneddon, WB; Bisello, A; Friedman, PA
Published in: J Biol Chem
April 13, 2018

G protein-coupled receptor (GPCR) signaling and trafficking are essential for cellular function and regulated by phosphorylation, β-arrestin, and ubiquitination. The GPCR parathyroid hormone receptor (PTHR) exhibits time-dependent reversible ubiquitination. The exact ubiquitination sites in PTHR are unknown, but they extend upstream of its intracellular tail. Here, using tandem MS, we identified Lys388 in the third loop and Lys484 in the C-terminal tail as primary ubiquitination sites in PTHR. We found that PTHR ubiquitination requires β-arrestin and does not display a preference for β-arrestin1 or -2. PTH stimulated PTHR phosphorylation at Thr387/Thr392 and within the Ser489-Ser493 region. Such phosphorylation events may recruit β-arrestin, and we observed that chemically or genetically blocking PTHR phosphorylation inhibits its ubiquitination. Specifically, Ala replacement at Thr387/Thr392 suppressed β-arrestin binding and inhibited PTHR ubiquitination, suggesting that PTHR phosphorylation and ubiquitination are interdependent. Of note, Lys-deficient PTHR mutants promoted normal cAMP formation, but exhibited differential mitogen-activated protein kinase (MAPK) signaling. Lys-deficient PTHR triggered early onset and delayed ERK1/2 signaling compared with wildtype PTHR. Moreover, ubiquitination of Lys388 and Lys484 in wildtype PTHR strongly decreased p38 signaling, whereas Lys-deficient PTHR retained signaling comparable to unstimulated wildtype PTHR. Lys-deficient, ubiquitination-refractory PTHR reduced cell proliferation and increased apoptosis. However, elimination of all 11 Lys residues in PTHR did not affect its internalization and recycling. These results pinpoint the ubiquitinated Lys residues in PTHR controlling MAPK signaling and cell proliferation and survival. Our findings suggest new opportunities for targeting PTHR ubiquitination to regulate MAPK signaling or manage PTHR-related disorders.

Duke Scholars

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Published In

J Biol Chem

DOI

EISSN

1083-351X

Publication Date

April 13, 2018

Volume

293

Issue

15

Start / End Page

5556 / 5571

Location

United States

Related Subject Headings

  • p38 Mitogen-Activated Protein Kinases
  • Ubiquitination
  • Receptor, Parathyroid Hormone, Type 1
  • Mutation, Missense
  • Mice
  • MAP Kinase Signaling System
  • Humans
  • HEK293 Cells
  • Cell Survival
  • Cell Proliferation
 

Citation

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Zhang, Q., Xiao, K., Liu, H., Song, L., McGarvey, J. C., Sneddon, W. B., … Friedman, P. A. (2018). Site-specific polyubiquitination differentially regulates parathyroid hormone receptor-initiated MAPK signaling and cell proliferation. J Biol Chem, 293(15), 5556–5571. https://doi.org/10.1074/jbc.RA118.001737
Zhang, Qiangmin, Kunhong Xiao, Hongda Liu, Lei Song, Jennifer C. McGarvey, W Bruce Sneddon, Alessandro Bisello, and Peter A. Friedman. “Site-specific polyubiquitination differentially regulates parathyroid hormone receptor-initiated MAPK signaling and cell proliferation.J Biol Chem 293, no. 15 (April 13, 2018): 5556–71. https://doi.org/10.1074/jbc.RA118.001737.
Zhang Q, Xiao K, Liu H, Song L, McGarvey JC, Sneddon WB, et al. Site-specific polyubiquitination differentially regulates parathyroid hormone receptor-initiated MAPK signaling and cell proliferation. J Biol Chem. 2018 Apr 13;293(15):5556–71.
Zhang, Qiangmin, et al. “Site-specific polyubiquitination differentially regulates parathyroid hormone receptor-initiated MAPK signaling and cell proliferation.J Biol Chem, vol. 293, no. 15, Apr. 2018, pp. 5556–71. Pubmed, doi:10.1074/jbc.RA118.001737.
Zhang Q, Xiao K, Liu H, Song L, McGarvey JC, Sneddon WB, Bisello A, Friedman PA. Site-specific polyubiquitination differentially regulates parathyroid hormone receptor-initiated MAPK signaling and cell proliferation. J Biol Chem. 2018 Apr 13;293(15):5556–5571.

Published In

J Biol Chem

DOI

EISSN

1083-351X

Publication Date

April 13, 2018

Volume

293

Issue

15

Start / End Page

5556 / 5571

Location

United States

Related Subject Headings

  • p38 Mitogen-Activated Protein Kinases
  • Ubiquitination
  • Receptor, Parathyroid Hormone, Type 1
  • Mutation, Missense
  • Mice
  • MAP Kinase Signaling System
  • Humans
  • HEK293 Cells
  • Cell Survival
  • Cell Proliferation