Negative regulation of Lck by Cbl ubiquitin ligase.
The Cbl-family ubiquitin ligases function as negative regulators of activated receptor tyrosine kinases by facilitating their ubiquitination and subsequent targeting to lysosomes. Cbl associates with the lymphoid-restricted nonreceptor tyrosine kinase Lck, but the functional relevance of this interaction remains unknown. Here, we demonstrate that T cell receptor and CD4 coligation on human T cells results in enhanced association between Cbl and Lck, together with Lck ubiquitination and degradation. A Cbl(-/-) T cell line showed a marked deficiency in Lck ubiquitination and increased levels of kinase-active Lck. Coexpression in 293T cells demonstrated that Lck kinase activity and Cbl ubiquitin ligase activity were essential for Lck ubiquitination and negative regulation of Lck-dependent serum response element-luciferase reporter activity. The Lck SH3 domain was pivotal for Cbl-Lck association and Cbl-mediated Lck degradation, with a smaller role for interactions mediated by the Cbl tyrosine kinase-binding domain. Finally, analysis of a ZAP-70-deficient T cell line revealed that Cbl inhibited Lck-dependent mitogen-activated protein kinase activation, and an intact Cbl RING finger domain was required for this functional effect. Our results demonstrate a direct, ubiquitination-dependent, negative regulatory role of Cbl for Lck in T cells, independent of Cbl-mediated regulation of ZAP-70.
Rao
Navin
N
Department of Medicine, Brigham and Women's Hospital, Harvard Medical School, Boston, MA 02115, USA.
Miyake
Sachiko
S
Reddi
Alagarsamy Lakku
AL
Douillard
Patrice
P
Ghosh
Amiya K
AK
Dodge
Ingrid L
IL
Zhou
Pengcheng
P
Fernandes
Norvin D
ND
Band
Hamid
H
D015704:CD4 Antigens / Q000378:metabolism; D002460:Cell Line; D004789:Enzyme Activation; D017353:Gene Deletion; D017930:Genes, Reporter / Q000235:genetics; D006801:Humans; D019169:Jurkat Cells; D008025:Ligases / Q000737:chemistry / Q000172:deficiency / Q000235:genetics / Q000378:metabolism*; D008213:Lymphocyte Activation; D019860:Lymphocyte Specific Protein Tyrosine Kinase p56(lck) / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D011485:Protein Binding; D011499:Protein Processing, Post-Translational; D017434:Protein Structure, Tertiary; D011505:Protein-Tyrosine Kinases / Q000378:metabolism; D011518:Proto-Oncogene Proteins / Q000737:chemistry / Q000172:deficiency / Q000235:genetics / Q000378:metabolism*; D050721:Proto-Oncogene Proteins c-cbl; D011948:Receptors, Antigen, T-Cell / Q000378:metabolism; D013601:T-Lymphocytes / Q000276:immunology / Q000378:metabolism; D025801:Ubiquitin / Q000378:metabolism*; D044767:Ubiquitin-Protein Ligases; D051746:ZAP-70 Protein-Tyrosine Kinase; D018909:src Homology Domains
D015704:CD4 Antigens; D011518:Proto-Oncogene Proteins; D011948:Receptors, Antigen, T-Cell; D025801:Ubiquitin; D050721:Proto-Oncogene Proteins c-cbl; D044767:Ubiquitin-Protein Ligases; D011505:Protein-Tyrosine Kinases; D019860:Lymphocyte Specific Protein Tyrosine Kinase p56(lck); D051746:ZAP-70 Protein-Tyrosine Kinase; C497261:ZAP70 protein, human; D008025:Ligases; C067419:CBL protein, human