β-Trcp mediates ubiquitination and degradation of the erythropoietin receptor and controls cell proliferation

Abstract

Control of intensity and duration of erythropoietin (Epo) signaling is necessary to tightly regulate red blood cell production. We have recently shown that the ubiquitin/ proteasome system plays a major role in the control of Epo-R signaling. Indeed, after Epo stimulation, Epo-R is ubiquitinated and its intracellular part is degraded by the proteasome, preventing further signal transduction. The remaining part of the receptor and associated Epo are internalized and degraded by the lysosomes. We show that ␤-Trcp is responsible for Epo-R ubiquitination and degradation. After Epo stimulation, ␤-Trcp binds to the Epo-R. This binding, like Epo-R ubiquitination, requires Jak2 activation. The Epo-R contains a typical DSG binding sequence for ␤-Trcp that is highly conserved among species. Interestingly, this sequence is located in a region of the Epo-R that is deleted in patients with familial polycythemia. Mutation of the serine residue of this motif to alanine IntroductionThe glycoprotein hormone erythropoietin (Epo) regulates the proliferation, survival, and differentiation of erythroid progenitor cells leading to the tight control of red blood cell production (reviewed in Lacombe and Mayeux 1 ). The Epo receptor (Epo-R) is a homodimer that belongs to the cytokine receptor superfamilly. Each Epo-R molecule of the dimer is constitutively associated with a Jak2 tyrosine kinase molecule that is absolutely required for Epo signaling. [2][3][4] Association of Jak2 with the Epo-R is also necessary for Epo-R maturation and cell surface expression. 5 Epo binding to its receptor induces a conformational change in the receptor complex that leads to Jak2 activation, subsequent phosphorylation of intracellular Epo-R Tyr residues, and activation of several intracellular signaling pathways, including Stat5, Ras/MAP kinase and PI3 kinase/Akt (reviewed by Damen et al 6 ). At the same time, desensitization processes are activated. The tyrosine phosphatase SHP-1 (SH2 domain-containing protein-tyrosine phosphatase-1) is recruited through phosphorylated Tyr 430 in human Epo-R and it dephosphorylates and deactivates Jak2. 7 CIS/SOCS family members are produced upon Epo stimulation and Cis, Socs-1, and Socs-3 suppress Epo signaling through their binding to the Epo-R complex. [8][9][10] Lastly, Epo and Epo-R are internalized and degraded. [11][12][13] Epo-induced Epo-R down-regulation is a complex process that involves Epo-R ubiquitination, proteasomes, and lysosomes. Epo-R is rapidly ubiquitinated at the cell surface after Epo binding. This process occurs before receptor internalization and requires Jak2 kinase activity. Most of the Epo-R intracellular domain is then degraded by proteasomes. This degradation process removes all the phosphorylated tyrosine residues in the intracellular domain, thereby preventing further signal transduction. The remaining part of the Epo/Epo-R complex is internalized and degraded by the lysosomes. 13 Thus, it appears that Epo-R ubiquitination plays a key role in Epo-R down-regulation during Epo stimulation...

Polyubiquitination of Prolactin Receptor Stimulates Its Internalization, Postinternalization Sorting, and Degradation via the Lysosomal Pathway

“…Remarkably, in similarity to PRLr, ␤-TrCP E3 ubiquitin ligase also plays a major role in regulating the stability of erythropoietin receptor (31) and growth hormone receptor (43). Although recruitment of ␤-TrCP to the latter receptor promotes its downregulation, the role of growth hormone receptor ubiquitination per se in this process remains to be elucidated.…”

Role of alternative phosphorylation and O-glycosylation of erythropoietinreceptor in modulating its function: an in silico study

“…The presence of proline at both +1 and −1 positions also supports its glycosylation potential. Glycosylation at Thr 423 might hinder β-TrCP binding, an enzyme involved in regulation of the EPO-R that ubiquitinates at K-428 and ultimately results in recycling EPO-R to the cell surface (Meyer et al, 2007). However, phosphorylation on the same residue inhibits the EPO-R recycling and decreases EPO sensitivity (Gross and Lodish, 2006).…”

Erythropoietin and Its Receptor in Breast Cancer: Putting Together the Pieces of the Puzzle

“…Analyzing the ubiquitination, internalization, and degradation model of Epo and its receptor in hematopoietic cells [41][42][43], at least two mechanisms may justify the confounding results obtained in breast cells and tissues. Let's put together the "scattered" pieces of Epo and EpoR findings in breast through (a) JAK-dependent ubiquitination/ degradation and (b) JAK-independent internalizationrecycling mechanisms.…”

“…In conjunction with 764 The Jigsaw Puzzle of Epo in Breast Cancer other kinases, JAK2 phosphorylates several tyrosine residues in the EpoR, creating docking sites for the SH2 domains of several signal transduction proteins (e.g., STAT5 and other signaling proteins become phosphorylated and activated, promote intracellular signaling, move to the nucleus, and activate gene expression) [33]. The JAK2-induced tyrosine-phosphorylated form of EpoR is rapidly ubiquitinated [44], an important step both for the proteasomal degradation of its intracellular domain (preventing further signal transduction) and for the targeting and routing of Epo and EpoR to lysosomes [41,43]. The proteasomal cleaved EpoR (still complexed with Epo protein) may be internalized (probably by a clathrin/caveolae-dependent mechanism) and may follow two fates: (a) lysosome targeting for the final proteolytic degradation of both Epo and cleaved EpoR [42] and (b) endosome formation for the extracellular secretion of the soluble form of cleaved EpoR and Epo.…”