JAK kinases control IL-5 receptor ubiquitination, degradation, and internalization.
IL-5, IL-3, and GM-CSF are related hematopoietic cytokines, which regulate the function of myeloid cells and are mediators of the allergic inflammatory response. These cytokines signal through heteromeric receptors containing a specific alpha chain and a shared signaling chain, betac. Previous studies demonstrated that the ubiquitin (Ub) proteasome degradation pathway was involved in signal termination of the betac-sharing receptors. In this study, the upstream molecular events leading to proteasome degradation of the IL-5 receptor (IL-5R) were examined. By using biochemical and flow cytometric methods, we show that JAK kinase activity is required for betac ubiquitination and proteasome degradation but only partially required for IL-5R internalization. Furthermore, we demonstrate the direct ubiquitination of the betac cytoplasmic domain and identify lysine residues 566 and 603 as sites of betac ubiquitination. Lastly, we show that ubiquitination of the betac cytoplasmic domain begins at the plasma membrane, increases after receptor internalization, and is degraded by the proteasome after IL-5R internalization. We propose an updated working model of IL-5R down-regulation, whereby IL-5 ligation of its receptor activates JAK2/1 kinases, resulting in betac tyrosine phosphorylation, ubiquitination, and IL-5R internalization. Once inside the cell, proteasomes degrade the betac cytoplasmic domain, and the truncated receptor complex is terminally degraded in the lysosomes. These data establish a critical role for JAK kinases and the Ub/proteasome degradation pathway in IL-5R down-regulation.
Martinez-Moczygemba
Margarita
M
Biology of Inflammation Center and Immunology, Allergy and Rheumatology Section, Department of Medicine, Baylor College of Medicine, One Baylor Plaza, BCM 285, Houston, TX 77030-3411, USA. mxm@bcm.tmc.edu
Huston
David P
DP
Lei
Jonathan T
JT
D002460:Cell Line; D002478:Cells, Cultured; D003593:Cytoplasm / Q000378:metabolism; D015536:Down-Regulation; D004705:Endocytosis; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004804:Eosinophils / Q000201:enzymology / Q000378:metabolism / Q000502:physiology; D006801:Humans; D053612:Janus Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism* / Q000502:physiology; D008239:Lysine / Q000737:chemistry; D008954:Models, Biological; D046988:Proteasome Endopeptidase Complex / Q000378:metabolism / Q000502:physiology; D053649:Receptors, Interleukin-5 / Q000378:metabolism*; D015398:Signal Transduction; D020032:Tyrphostins / Q000494:pharmacology; D025801:Ubiquitin / Q000378:metabolism*; D019061:src-Family Kinases / Q000378:metabolism*
D004791:Enzyme Inhibitors; D053649:Receptors, Interleukin-5; D020032:Tyrphostins; D025801:Ubiquitin; C095512:alpha-cyano-(3,4-dihydroxy)-N-benzylcinnamide; D053612:Janus Kinases; C060902:lyn protein-tyrosine kinase; D019061:src-Family Kinases; D046988:Proteasome Endopeptidase Complex; D008239:Lysine