Newly synthesised major histocompatibility complex class II molecules associate with invariant chains (Ii) to form nonameric complexes. These complexes are transported to endosomes, where proteolytic enzymes generate alphabeta class II dimers associated with nested Ii-derived peptides. These peptides are then exchanged with antigen peptide, and mature class II molecules reach the cell surface. The role of the actin cytoskeleton in the transport and maturation of class II molecules has not been studied. We show here that upon treatment with cytochalasin D (cyto D), the rate of Ii degradation is drastically reduced in B cells. Cyto D treatment also leads to a delayed appearance of stable forms of class II molecules, and a reduced presentation efficiency of antigen determinants requiring newly synthesised class II molecules. Under such conditions, we found that invariant chain fragments and class II molecules are accumulated in early and late endosomal compartments, whereas the leupeptin protease inhibitor induces their accumulation in lysosomal compartments. The addition of cyto D to leupeptin blocks the delivery of class II/invariant chain complexes to lysosomes, and further inhibits degradation of Ii. The dynamics of the actin cytoskeleton can therefore control the meeting point between newly synthesised class II molecules and lysosomal proteases, involved in Ii degradation and antigen peptide loading.

REFERENCES

Adorini
L.
,
Ullrich
S. J.
,
Appella
E.
,
Fuchs
S.
(
1990
).
Inhibition by brefeldin A of presentation of exogenous protein antigens to MHC class II-restricted T cells.
Nature
346
,
63
66
Allen
P. M.
,
Unanue
E. R.
(
1984
).
Differential requirements for antigen processing by macrophages for lysozyme-specific T cell hybridomas.
J. Immunol
132
,
1077
1079
Amigorena
S.
,
Webster
P.
,
Drake
J.
,
Newcomb
J.
,
Cresswell
P.
,
Mellman
I.
(
1995
).
Invariant chain cleavage and peptide loading in major histocompatibility complex class II vesicles.
J. Exp. Med
181
,
1729
1741
Arneson
L. S.
,
Miller
J.
(
1995
).
Efficient endosomal localization of major histocompatibility complex class II-invariant chain complexes requires multimerization of the invariant chain targeting sequence.
J. Cell Biol
129
,
1217
1228
Barois
N.
,
Forquet
F.
,
Davoust
J.
(
1997
).
Selective modulation of the major histocompatibility complex class II antigen presentation pathway following B cell receptor ligation and protein kinase C activation.
J. Biol. Chem
272
,
3641
3647
Bertolino
P.
,
Rabourdin-Combe
C.
(
1996
).
The MHC class II-associated invariant chain: A molecule with multiple roles in MHC class II biosynthesis and antigen presentation to CD4+T cells.
Crit. Rev. Immunol
16
,
359
379
Brachet
V.
,
Raposo
G.
,
Amigorena
S.
,
Mellman
I.
(
1997
).
Ii chain controls the transport of major histocompatibility complex class II molecules to and from lysosomes.
J. Cell Biol
137
,
51
65
Cella
M.
,
Engering
A.
,
Pinet
V.
,
Pieters
J.
,
Lanzavecchia
A.
(
1997
).
Inflammatory stimuli induce accumulation of MHC class II complexes on dendritic cells.
Nature
388
,
782
787
Denzin
L. K.
,
Cresswell
P.
(
1995
).
HLA-DM induces CLIP dissociation from MHC class IIdimers and facilitates peptide loading.
Cell
82
,
155
165
Durrbach
A.
,
Louvard
D.
,
Coudrier
E.
(
1996
).
Actin filaments facilitate two steps of endocytosis.
J. Cell Sci
109
,
457
465
Fernandez-Borja
M.
,
Verwoerd
D.
,
Sanderson
F.
,
Aerts
H.
,
Trowsdale
J.
,
Tulp
A.
,
Neefjes
J.
(
1996
).
HLA-DM and MHC class II molecules co-distribute with peptidase-containing lysosomal subcompartments.
Int. Immunol
8
,
625
640
Germain
R. N.
,
Hendrix
L. R.
(
1991
).
MHC class II structure, occupancy and surface expression determined by post-endoplasmic reticulum antigen binding.
Nature
353
,
134
139
Germain
R. N.
(
1994
).
MHC-dependent antigen processing and peptide presentation: Providing ligands for T lymphocyte activation.
Cell
76
,
287
299
Ghosh
P.
,
Amaya
M.
,
Mellins
E.
,
Wiley
D. C.
(
1995
).
The structure of an intermediate in class II MHC maturation: CLIP bound to HLA-DR3.
Nature
378
,
457
462
Gottlieb
T. A.
,
Ivanov
I. E.
,
Adesnik
M.
,
Sabatini
D. D.
(
1993
).
Actin microfilaments play a critical role in endocytosis at the apical but not the basolateral surface of polarized epithelial cells.
J. Cell Biol
120
,
695
710
Granger
B. L.
,
Green
S. A.
,
Gabel
C. A.
,
Howe
C. L.
,
Mellman
I.
,
Helenius
A.
(
1990
).
Characterization and cloning of lgp 110, a lysosomal membrane glycoprotein from mouse and rat cells.
J. Biol. Chem
265
,
12036
12043
Griffin
J. P.
,
Chu
R.
,
Harding
C. V.
(
1997
).
Early endosomes and a late endocytic compartment generate different peptide-class II MHC complexes via distinct processing mechanisms.
J. Immunol
158
,
1523
1532
Griffiths
G.
,
Hoflack
B.
,
Simons
K.
,
Mellman
I.
,
Kornfeld
S.
(
1988
).
The mannose 6-phosphate receptor and the biogenesis of lysosomes.
Cell
52
,
329
341
Harding
C. V.
,
Roof
R. W.
,
Unanue
E. R.
(
1989
).
Turnover of Ia-peptide complexes is facilitated in viable antigen-presenting cells: Biosynthetic turnover of Ia vs. peptide exchange.
Proc. Nat. Acad. Sci. USA
86
,
4230
4234
Liu
K.-J.
,
Parikh
V. S.
,
Tucker
P. W.
,
Kim
B. S.
(
1994
).
Surface immunoglobulins mediate efficient transport of antigen to lysosomal compartments resulting in enhanced specific antigen presentation by B cells.
Eur. J. Immunol
24
,
2755
2760
Long
E. O.
,
LaVaute
T.
,
Pinet
V.
,
Jaraquemada
D.
(
1994
).
Invariant chain prevents the HLA-DR-restricted presentation of a cytosolic peptide.
J. Immunol
153
,
1487
1494
Lorenz
R. G.
,
Tyler
A. N.
,
Allen
P. M.
(
1988
).
T cell recognition of bovine ribonuclease: Self/non-self discrimination at the level of binding to the I-Akmolecule.
J. Immunol
141
,
4124
4128
MacLean-Fletcher
S.
,
Pollard
T. D.
(
1980
).
Mechanisms of action of cyto B on actin.
Cell
20
,
329
341
Maric
M. A.
,
Taylor
M. D.
,
Blum
J. S.
(
1994
).
Endosomal aspartic proteinases are required for invariant-chain processing.
Proc. Nat. Acad. Sci. USA
91
,
2171
2175
Melamed
I.
,
Downey
G. P.
,
Aktories
K.
,
Roifman
C. M.
(
1991
).
Microfilament assembly is required for antigen-receptor-mediated activation of human B lymphocytes.
J. Immunol
147
,
1139
1146
Mitchell
R. N.
,
Barnes
K. A.
,
Grupp
S. A.
,
Sanchez
M.
,
Misulovin
Z.
,
Nussenzweig
M. C.
,
Abbas
A. K.
(
1995
).
Intracellular targeting of antigens internalized by membrane immunoglobulin in B lymphocytes.
J. Exp. Med
181
,
1705
1714
Morkowski
S.
,
Goldrath
A. W.
,
Eastman
S.
,
Ramachandra
L.
,
Freed
D. C.
,
Whiteley
P.
,
Rudensky
A. Y.
(
1995
).
T cell recognition of major histocompatibility complex class II complexes with invariant chain processing intermediates.
J. Exp. Med
182
,
1403
1413
Nadimi
F.
,
Moreno
J.
,
Momburg
F.
,
Heuser
A.
,
Fuchs
S.
,
Adorini
L.
,
Hämmerling
G. J.
(
1991
).
Antigen presentation of hen egg-white lysozyme but not of ribonuclease A is augmented by the major histocompatibility complex class II-associated invariant chain.
Eur. J. Immunol
21
,
1255
1263
Neefjes
J. J.
,
Ploegh
H. L.
(
1992
).
Inhibition of endosomal proteolytic activity by leupeptin blocks surface expression of MHC class II molecules and their conversion to SDS resistantheterodimers in endosomes.
EMBO J
11
,
411
416
Newell
M. K.
,
Justement
L. B.
,
Miles
C. R.
,
Freed
J. H.
(
1988
).
Biochemical characterization of proteins that co-purify with class II antigens of the murine MHC.
J. Immunol
140
,
1930
1938
Nguyen
Q. V.
,
Knapp
W.
,
Humphreys
R. E.
(
1989
).
Inhibition by leupeptin and antipain of the intracellular proteolysis of Ii.
Hum. Immunol
24
,
153
163
Odorizzi
C. G.
,
Trowbridge
I. S.
,
Xue
L.
,
Hopkins
C. R.
,
Davis
C. D.
,
Collawn
J. F.
(
1994
).
Sorting signals in the MHC class II invariant chain cytoplasmic tail and transmembrane region determine trafficking to an endocytic processing compartment.
J. Cell Biol
126
,
317
330
Pierre
P.
,
Turley
S. J.
,
Gatti
E.
,
Hull
M.
,
Meltzer
J.
,
Mirza
A.
,
Inaba
K.
,
Steinman
R. M.
,
Mellman
I.
(
1997
).
Developmental regulation of MHC class II transport in mouse dendritic cells.
Nature
388
,
787
792
Pieters
J.
,
Bakke
O.
,
Dobberstein
B.
(
1993
).
The MHC class II-associated invariant chain contains two endosomal targeting signals within its cytoplasmic tail.
J. Cell Sci
106
,
831
846
Pinet
V.
,
Vergelli
M.
,
Martin
R.
,
Bakke
O.
,
Long
E. O.
(
1995
).
Antigen presentation mediated by recycling of surface HLA-DR molecules.
Nature
375
,
603
606
Pond
L.
,
Watts
C.
(
1997
).
Characterization of transport of newly assembled, T cell-stimolatory MHC class II-peptide complexes from MHC class II compartments to the cell surface.
J. Immunol
159
,
543
553
Qiu
Y.
,
Xu
X.
,
Wandinger-Ness
A.
,
Dalke
D. P.
,
Pierce
S. K.
(
1994
).
Separation of subcellular compartments containing distinct functional forms of MHC class II.
J. Cell Biol
125
,
595
605
Riese
R. J.
,
Wolf
P. R.
,
Brömme
D.
,
Natkin
L. R.
,
Villadangos
J. A.
,
Ploegh
H. L.
,
Chapman
H. A.
(
1996
).
Essential role for cathepsin S in MHC class II-associated invariant chain processing and peptide loading.
Immunity
4
,
357
366
Roche
P. A.
,
Cresswell
P.
(
1991
).
Proteolysis of the class II-associated invariant chain generates a peptide binding site in intracellular HLA-DR molecules.
Proc. Nat. Acad. Sci. USA
88
,
3150
3154
Roche
P. A.
,
Marks
M. S.
,
Cresswell
P.
(
1991
).
Formation of a nine-subunit complex by HLA class II glycoproteins and the invariant chain.
Nature
354
,
392
394
Romagnoli
P.
,
Germain
R. N.
(
1994
).
The CLIP region of invariant chain plays a critical role in regulating major histocompatibility complex class II folding, transport, and peptide occupancy.
J. Exp. Med
180
,
1107
1113
Salamero
J.
,
Humbert
M.
,
Cosson
P.
,
Davoust
J.
(
1990
).
Mouse B lymphocyte specific endocytosis and recycling of MHC class II molecules.
EMBO J
9
,
3489
3496
Sanderson
F.
,
Kleijmeer
M. J.
,
Kelly
A.
,
Verwoerd
D.
,
Tulp
A.
,
Neefjes
J. J.
,
Geuze
H. J.
,
Trowsdale
J.
(
1994
).
Accumulation of HLA-DM, a regulator of antigen presentation, in MHC class II compartments.
Science
266
,
1566
1569
Sanderson
F.
,
Thomas
C.
,
Neefjes
J.
,
Trowsdale
J.
(
1996
).
Association between HLA-DM and HLA-DR in vivo.
Immunity
4
,
87
96
Soreng
K. M.
,
Moore
J. C.
,
Sherman
M. A.
,
Jensen
P. E.
(
1994
).
Requirement for protein synthesis in antigen processing by B cells.
Cell. Immunol
157
,
277
290
Soreng
K. M.
,
Weber
D. A.
,
joshi
H.
,
Moore
J. C.
,
Jensen
P. E.
(
1995
).
A role for microfilaments but not microtubules in processing soluble antigens.
Cell. Immunol
166
,
25
34
St-Pierre
Y.
,
Watts
T. H.
(
1990
).
MHC class II-restricted presentation of native protein antigen by B cells is inhibitable by cycloheximide and brefeldin A.
J. Immunol
145
,
812
818
Tulp
A.
,
Verwoerd
D.
,
Dobberstein
B.
,
Ploegh
H. L.
,
Pieters
J.
(
1994
).
Isolation and characterization of the intracellular MHC class II compartment.
Nature
369
,
120
126
Wade
W. F.
,
Chen
Z. Z.
,
Maki
R.
,
McKercher
S.
,
Palmer
E.
,
Cambier
J. C.
,
Freed
J. H.
(
1989
).
Altered I-A protein-mediated transmembrane signaling in B cells that express truncated I-Akprotein.
Proc. Nat. Acad. Sci. USA
86
,
6297
6301
Wade
W. F.
,
Freed
J. H.
,
Edidin
M.
(
1989
).
Translational diffusion of class II major histocompatibility complex molecules is constrained by their cytoplasmic domains.
J. Cell Biol
109
,
3325
3331
West
M. A.
,
Lucocq
J. M.
,
Watts
C.
(
1994
).
Antigen processing and class II MHC peptide-loading compartments in human B-lymphoblastoid cells.
Nature
369
,
147
151
Winzler
C.
,
Rovere
P.
,
Rescigno
M.
,
Granucci
F.
,
Penna
G.
,
Adorini
L.
,
Zimmermann
V. S.
,
Davoust
J.
,
Ricciardi-Castagnoli
P.
(
1997
).
Maturation stages of mouse dendritic cells in growth factor-dependent long-term cultures.
J. Exp. Med
185
,
317
328
Wubbolts
R.
,
Fernandez-Borja
M.
,
Oomen
L.
,
Verwoerd
D.
,
Janssen
H.
,
Calafat
J.
,
Tulp
A.
,
Dusseljee
S.
,
Neefjes
J.
(
1996
).
Direct vesicular transport of MHC class II molecules from lysosomal structures to the cell surface.
J. Cell Biol
135
,
611
622
Xu
M.
,
Capraro
G. A.
,
Daibata
M.
,
Reyes
V. E.
,
Humphreys
R. E.
(
1994
).
Cathepsin B cleavage and release of invariant chain from MHC class II molecules follow a staged pattern.
Mol. Immunol
31
,
723
731
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