Activatory and inhibitory Fcy receptors augment rituximab-mediated internalization of CD20 independent of signaling via the cytoplasmic domain
Activatory and inhibitory Fcy receptors augment rituximab-mediated internalization of CD20 independent of signaling via the cytoplasmic domain
Type I anti-CD20 mAb such as rituximab and ofatumumab engage with the inhibitory Fc?R, Fc?RIIb on the surface of B cells, resulting in immunoreceptor tyrosine-based inhibitory motif (ITIM) phosphorylation. Internalization of the CD20·mAb·Fc?RIIb complex follows, the rate of which correlates with Fc?RIIb expression. In contrast, although type II anti-CD20 mAb such as tositumomab and obinutuzumab also interact with and activate Fc?RIIb, this interaction fails to augment the rate of CD20·mAb internalization, raising the question of whether ITIM phosphorylation plays any role in this process. We have assessed the molecular requirements for the internalization process and demonstrate that in contrast to internalization of IgG immune complexes, Fc?RIIb-augmented internalization of rituximab-ligated CD20 occurs independently of the Fc?RIIb ITIM, indicating that signaling downstream of Fc?RIIb is not required. In transfected cells, activatory Fc?RI, Fc?RIIa, and Fc?RIIIa augmented internalization of rituximab-ligated CD20 in a similar manner. However, Fc?RIIa mediated a slower rate of internalization than cells expressing equivalent levels of the highly homologous Fc?RIIb. The difference was maintained in cells expressing Fc?RIIa and Fc?RIIb lacking cytoplasmic domains and in which the transmembrane domains had been exchanged. This difference may be due to increased degradation of Fc?RIIa, which traffics to lysosomes independently of rituximab. We conclude that the cytoplasmic domain of Fc?R is not required for promoting internalization of rituximab-ligated CD20. Instead, we propose that Fc?R provides a structural role in augmenting endocytosis that differs from that employed during the endocytosis of immune complexes.
5424-5437
Vaughan, A.T.
bfb2ceab-a592-457e-89f9-00fcd1dddbdb
Chan, C.H.T.
b109c93f-7e9a-44ee-ad12-da757b1b11fc
Klein, C.
4546582b-512e-4725-b2ca-c160b1a967cd
Glennie, M.J.
9f6f0eff-4560-48c2-80cd-0ec116110ded
Beers, S.A.
a02548be-3ffd-41ab-9db8-d6e8c3b499a2
Cragg, M.
ec97f80e-f3c8-49b7-a960-20dff648b78c
27 February 2015
Vaughan, A.T.
bfb2ceab-a592-457e-89f9-00fcd1dddbdb
Chan, C.H.T.
b109c93f-7e9a-44ee-ad12-da757b1b11fc
Klein, C.
4546582b-512e-4725-b2ca-c160b1a967cd
Glennie, M.J.
9f6f0eff-4560-48c2-80cd-0ec116110ded
Beers, S.A.
a02548be-3ffd-41ab-9db8-d6e8c3b499a2
Cragg, M.
ec97f80e-f3c8-49b7-a960-20dff648b78c
Vaughan, A.T., Chan, C.H.T., Klein, C., Glennie, M.J., Beers, S.A. and Cragg, M.
(2015)
Activatory and inhibitory Fcy receptors augment rituximab-mediated internalization of CD20 independent of signaling via the cytoplasmic domain.
The Journal of Biological Chemistry, 290, .
(doi:10.1074/jbc.M114.593806).
(PMID:25568316)
Abstract
Type I anti-CD20 mAb such as rituximab and ofatumumab engage with the inhibitory Fc?R, Fc?RIIb on the surface of B cells, resulting in immunoreceptor tyrosine-based inhibitory motif (ITIM) phosphorylation. Internalization of the CD20·mAb·Fc?RIIb complex follows, the rate of which correlates with Fc?RIIb expression. In contrast, although type II anti-CD20 mAb such as tositumomab and obinutuzumab also interact with and activate Fc?RIIb, this interaction fails to augment the rate of CD20·mAb internalization, raising the question of whether ITIM phosphorylation plays any role in this process. We have assessed the molecular requirements for the internalization process and demonstrate that in contrast to internalization of IgG immune complexes, Fc?RIIb-augmented internalization of rituximab-ligated CD20 occurs independently of the Fc?RIIb ITIM, indicating that signaling downstream of Fc?RIIb is not required. In transfected cells, activatory Fc?RI, Fc?RIIa, and Fc?RIIIa augmented internalization of rituximab-ligated CD20 in a similar manner. However, Fc?RIIa mediated a slower rate of internalization than cells expressing equivalent levels of the highly homologous Fc?RIIb. The difference was maintained in cells expressing Fc?RIIa and Fc?RIIb lacking cytoplasmic domains and in which the transmembrane domains had been exchanged. This difference may be due to increased degradation of Fc?RIIa, which traffics to lysosomes independently of rituximab. We conclude that the cytoplasmic domain of Fc?R is not required for promoting internalization of rituximab-ligated CD20. Instead, we propose that Fc?R provides a structural role in augmenting endocytosis that differs from that employed during the endocytosis of immune complexes.
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Accepted/In Press date: 24 December 2014
e-pub ahead of print date: 7 January 2015
Published date: 27 February 2015
Organisations:
Cancer Sciences
Identifiers
Local EPrints ID: 390377
URI: http://eprints.soton.ac.uk/id/eprint/390377
ISSN: 0021-9258
PURE UUID: a6645acc-3b78-45c9-8659-a1a62a12a2a3
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Date deposited: 01 Apr 2016 07:58
Last modified: 15 Mar 2024 03:08
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Author:
A.T. Vaughan
Author:
C.H.T. Chan
Author:
C. Klein
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