Efficient internalization of MHC I requires lysine-11 and lysine-63 mixed linkage polyubiquitin chains.
The downregulation of cell surface receptors by endocytosis is a fundamental requirement for the termination of signalling responses and ubiquitination is a critical regulatory step in receptor regulation. The K5 gene product of Kaposi's sarcoma-associated herpesvirus is an E3 ligase that ubiquitinates and downregulates several cell surface immunoreceptors, including major histocompatibility complex (MHC) class I molecules. Here, we show that K5 targets the membrane proximal lysine of MHC I for conjugation with mixed linkage polyubiquitin chains. Quantitative mass spectrometry revealed an increase in lysine-11, as well as lysine-63, linked polyubiquitin chains on MHC I in K5-expressing cells. Using a combination of mutant ubiquitins and MHC I molecules expressing a single cytosolic lysine residue, we confirm a functional role for lysines-11 and -63 in K5-mediated MHC I endocytosis. We show that lysine-11 linkages are important for receptor endocytosis, and that complex mixed linkage polyubiquitin chains are generated in vivo.
Boname
Jessica M
JM
School of Clinical Medicine, Cambridge Institute for Medical Research, University of Cambridge, Cambridge, CB2 0XY, UK.
Thomas
Mair
M
Stagg
Helen R
HR
Xu
Ping
P
Peng
Junmin
J
Lehner
Paul J
PJ
D000595:Amino Acid Sequence; D004705:Endocytosis / Q000502:physiology*; D008040:Genetic Linkage; D006367:HeLa Cells; D006801:Humans; D008239:Lysine / Q000378:metabolism*; D008285:Major Histocompatibility Complex / Q000502:physiology*; D013058:Mass Spectrometry; D008969:Molecular Sequence Data; D009154:Mutation; D025821:Polyubiquitin / Q000737:chemistry / Q000378:metabolism*; D011956:Receptors, Cell Surface; D015398:Signal Transduction; D044767:Ubiquitin-Protein Ligases / Q000378:metabolism; D054875:Ubiquitination
D011956:Receptors, Cell Surface; D025821:Polyubiquitin; D044767:Ubiquitin-Protein Ligases; D008239:Lysine