An LI and ML motif in the cytoplasmic tail of the MHC-associated invariant chain mediate rapid internalization

1994 ◽  
Vol 107 (7) ◽  
pp. 2021-2032 ◽  
Author(s):  
B. Bremnes ◽  
T. Madsen ◽  
M. Gedde-Dahl ◽  
O. Bakke
Keyword(s):  
Plasma Membrane ◽  
Mhc Class Ii ◽  
Transmembrane Protein ◽  
Point Mutations ◽  
Cytoplasmic Tail ◽  
Class Ii ◽  
Endocytic Pathway ◽  
Invariant Chain ◽  
Chimeric Proteins ◽  
Endosomal Pathway

Invariant chain (Ii) is a transmembrane protein that associates with the MHC class II molecules in the endoplasmic reticulum. Two regions of the 30 residue cytoplasmic tail of Ii contain sorting information able to direct Ii to the endocytic pathway. The full-length cytoplasmic tail of Ii and the two tail regions were fused to neuraminidase (NA) forming chimeric proteins (INA). Ii is known to form trimers and when INA was transfected into COS cells it assembled as a tetramer like NA. The INA molecules were targeted to the endosomal pathway and cotransfection with Ii showed that both molecules appeared in the same vesicles. By labelling the INA fusion proteins with iodinated antibody it was found that molecules with either endocytosis signal were expressed at the plasma membrane and internalized rapidly. Point mutations revealed that an LI motif within the first region of the cytoplasmic tail and an ML motif in the second region were essential for efficient internalization. The region containing the LI motif is required for Ii to induce large endosomes but a functional LI internalization motif was not fundamental for this property. The cytoplasmic tail of Ii is essential for efficient targeting of the class II molecules to endosomes and the dual LI and ML motif may thus be responsible for directing these molecules to the endosomal pathway, possibly via the plasma membrane.

scholarly journals The MHC class II-associated invariant chain contains two endosomal targeting signals within its cytoplasmic tail

1993 ◽  
Vol 106 (3) ◽  
pp. 831-846 ◽  
Author(s):  
J. Pieters ◽  
O. Bakke ◽  
B. Dobberstein
Keyword(s):  
Plasma Membrane ◽  
Amino Acid ◽  
Mhc Class Ii ◽  
Cytoplasmic Tail ◽  
Class Ii ◽  
Endocytic Pathway ◽  
Invariant Chain ◽  
Amino Acid Residues ◽  
Marker Proteins ◽  
Membrane Spanning

The oligomeric complex formed by major histocompatibility complex (MHC) class II alpha and beta chains and invariant chain (Ii) assembles in the endoplasmic reticulum and is then transported via the Golgi complex to compartments of the endocytic pathway. When Ii alone is expressed in CV1 cells it is sorted to endosomes. The Ii cytoplasmic tail has been found to be essential for targeting to these compartments. In order to characterize further the signals responsible for endosomal targeting, we have deleted various segments of the cytoplasmic tail. The Ii mutants were transiently expressed and the cellular location of the proteins was analyzed biochemically and morphologically. The cytoplasmic tail of Ii was found to contain two endosomal targeting sequences within its cytoplasmic tail; one targeting sequence was present within amino acid residues 12–29 and deletion of this segment revealed the presence of a second endosomal targeting sequence, located within the first 11 amino acid residues. The presence of a leucine-isoleucine pair at positions 7 and 8 within this sequence was found to be essential for endosomal targeting. In addition, the presence of this L-I motif lead to accumulation of Ii molecules in large endosomal vacuoles containing lysosomal marker proteins. Both wild type Ii and Ii mutant molecules containing only one endosomal targeting sequence were rapidly internalized from the plasma membrane. When the Ii cytoplasmic tail was fused to the membrane-spanning region of neuraminidase, a resident plasma membrane protein, the resulting chimera (INA) was found in endocytic compartments containing lysosomal marker proteins. Thus the cytoplasmic tail of Ii is sufficient for targeting to the endocytic/lysosomal pathway.

scholarly journals Sorting signals in the MHC class II invariant chain cytoplasmic tail and transmembrane region determine trafficking to an endocytic processing compartment.

1994 ◽  
Vol 126 (2) ◽  
pp. 317-330 ◽  
Author(s):  
C G Odorizzi ◽  
I S Trowbridge ◽  
L Xue ◽  
C R Hopkins ◽  
C D Davis ◽  
...  
Keyword(s):  
Plasma Membrane ◽  
Mhc Class Ii ◽  
Cytoplasmic Tail ◽  
Class Ii ◽  
Endocytic Pathway ◽  
Invariant Chain ◽  
Transmembrane Region ◽  
Coated Pits ◽  
Endocytic Compartment ◽  
Sorting Signals

Targeting of MHC class II molecules to the endocytic compartment where they encounter processed antigen is determined by the invariant chain (Ii). By analysis of Ii-transferrin receptor (TR) chimera trafficking, we have identified sorting signals in the Ii cytoplasmic tail and transmembrane region that mediate this process. Two non-tyrosine-based sorting signals in the Ii cytoplasmic tail were identified that mediate localization to plasma membrane clathrin-coated pits and promote rapid endocytosis. Leu7 and Ile8 were required for the activity of the signal most distal to the cell membrane whereas Pro15 Met16 Leu17 were important for the membrane-proximal signal. The same or overlapping non-tyrosine-based sorting signals are essential for delivery of Ii-TR chimeras, either by an intracellular route or via the plasma membrane, to an endocytic compartment where they are rapidly degraded. The Ii transmembrane region is also required for efficient delivery to this endocytic processing compartment and contains a signal distinct from the Ii cytoplasmic tail. More than 80% of the Ii-TR chimera containing the Ii cytoplasmic tail and transmembrane region is delivered directly to the endocytic pathway by an intracellular route, implying that the Ii sorting signals are efficiently recognized by sorting machinery located in the trans-Golgi.

scholarly journals Relationship between invariant chain expression and major histocompatibility complex class II transport into early and late endocytic compartments.

1993 ◽  
Vol 177 (3) ◽  
pp. 583-596 ◽  
Author(s):  
P Romagnoli ◽  
C Layet ◽  
J Yewdell ◽  
O Bakke ◽  
R N Germain
Keyword(s):  
Major Histocompatibility Complex ◽  
Mhc Class Ii ◽  
Class Ii ◽  
Endocytic Pathway ◽  
Invariant Chain ◽  
Major Histocompatibility ◽  
Histocompatibility Complex ◽  
Late Endosomes ◽  
Early Endosomes ◽  
Endocytic Compartments

Invariant chain (Ii), which associates with major histocompatibility complex (MHC) class II molecules in the endoplasmic reticulum, contains a targeting signal for transport to intracellular vesicles in the endocytic pathway. The characteristics of the target vesicles and the relationship between Ii structure and class II localization in distinct endosomal subcompartments have not been well defined. We demonstrate here that in transiently transfected COS cells expressing high levels of the p31 or p41 forms of Ii, uncleaved Ii is transported to and accumulates in transferrin-accessible (early) endosomes. Coexpressed MHC class II is also found in this same compartment. These early endosomes show altered morphology and a slower rate of content movement to later parts of the endocytic pathway. At more moderate levels of Ii expression, or after removal of a highly conserved region in the cytoplasmic tail of Ii, coexpressed class II molecules are found primarily in vesicles with the characteristics of late endosomes/prelysosomes. The Ii chains in these late endocytic vesicles have undergone proteolytic cleavage in the lumenal region postulated to control MHC class II peptide binding. These data indicate that the association of class II with Ii results in initial movement to early endosomes. At high levels of Ii expression, egress to later endocytic compartments is delayed and class II-Ii complexes accumulate together with endocytosed material. At lower levels of Ii expression, class II-Ii complexes are found primarily in late endosomes/prelysosomes. These data provide evidence that the route of class II transport to the site of antigen processing and loading involves movement through early endosomes to late endosomes/prelysosomes. Our results also reveal an unexpected ability of intact Ii to modify the structure and function of the early endosomal compartment, which may play a role in regulating this processing pathway.

scholarly journals Related Leucine-based Cytoplasmic Targeting Signals in Invariant Chain and Major Histocompatibility Complex Class II Molecules Control Endocytic Presentation of Distinct Determinants in a Single Protein

1997 ◽  
Vol 185 (3) ◽  
pp. 429-438 ◽  
Author(s):  
Guangming Zhong ◽  
Paola Romagnoli ◽  
Ronald N. Germain
Keyword(s):  
Major Histocompatibility Complex ◽  
Major Histocompatibility Complex Class ◽  
Cytoplasmic Tail ◽  
Class Ii ◽  
Endocytic Pathway ◽  
Invariant Chain ◽  
Antigenic Peptides ◽  
Complex Class ◽  
Major Histocompatibility ◽  
Histocompatibility Complex

Leucine-based signals in the cytoplasmic tail of invariant chain (Ii) control targeting of newly synthesized major histocompatibility complex class II molecules to the endocytic pathway for acquisition of antigenic peptides. Some protein determinants, however, do not require Ii for effective class II presentation, although endocytic processing is still necessary. Here we demonstrate that a dileucine-based signal in the cytoplasmic tail of the class II β chain is critical for this Ii-independent presentation. Elimination or mutation of this signal reduces the rate of re-entry of mature surface class II molecules into the endocytic pathway. Antigen presentation controlled by this signal does not require newly synthesized class II molecules and appears to involve determinants requiring only limited proteolysis for exposure, whereas the opposite is true for Ii-dependent determinants. This demonstrates that related leucine-based trafficking signals in Ii and class II control the functional presentation of protein determinants with distinct processing requirements, suggesting that the peptide binding sites of newly synthesized versus mature class II molecules are made available for antigen binding in distinct endocytic compartments under the control of these homologous cytoplasmic signals. This permits capture of protein fragments produced optimally under distinct conditions of pH and proteolytic activity.

scholarly journals A block in degradation of MHC class II-associated invariant chain correlates with a reduction in transport from endosome carrier vesicles to the prelysosome compartment

1992 ◽  
Vol 103 (3) ◽  
pp. 811-822 ◽  
Author(s):  
S. Zachgo ◽  
B. Dobberstein ◽  
G. Griffiths
Keyword(s):  
Mhc Class Ii ◽  
Antigen Processing ◽  
Intracellular Transport ◽  
Protein Transport ◽  
Multivesicular Body ◽  
Class Ii ◽  
Endocytic Pathway ◽  
Invariant Chain ◽  
Intermediate Form ◽  
General Effect

Invariant chain (Ii) associated with MHC class II molecule is processed proteolytically via several distinct intermediates during its intracellular transport through endosomal compartments. Leupeptin added to the culture medium blocks processing of Ii, prevents its dissociation from the class II molecules and leads to an intracellular accumulation of a 22 kDa intermediate form of Ii. We show here that leupeptin has a very general effect on protein transport in the endocytic pathway. When added to Mel Juso cells leupeptin reduces the transport of endocytosed material from multivesicular body-like, endosome carrier vesicles (ECV) to the prelysosomal compartment (late endosome) and leads to a concomitant increase in the number of ECV. Our results argue that one effect of leupeptin, related to antigen processing and presentation, is to block transport of antigen and/or MHC class II molecules to prelysosomal compartments.

scholarly journals Epitope-specific enhancement of antigen presentation by invariant chain.

1993 ◽  
Vol 178 (4) ◽  
pp. 1453-1458 ◽  
Author(s):  
F Momburg ◽  
S Fuchs ◽  
J Drexler ◽  
R Busch ◽  
M Post ◽  
...  
Keyword(s):  
Antigen Presentation ◽  
Mhc Class Ii ◽  
Brefeldin A ◽  
Class Ii ◽  
Endocytic Pathway ◽  
Invariant Chain ◽  
Target Class ◽  
Egg White Lysozyme ◽  
Hen Egg White ◽  
Enhance Antigen Presentation

The MHC class II-associated invariant chain (Ii) is involved in the intracellular sorting of class II molecules to the endocytic pathway where peptides from processed exogenous antigens are bound, and thereby Ii is thought to enhance antigen presentation. Here we demonstrate that presentation of only one out of five epitopes of a given antigen is augmented by Ii. We have compared the presentation of five different epitopes derived from hen egg white lysozyme (HEL) to Ak-restricted T hybridomas by rat-2 fibroblasts transfected with A alpha k and A beta k (RKK) and RKK cells supertransfected with the mouse invariant chain (RKKI). Only the presentation of the HEL epitope 46-61 was enhanced whereas the presentation of the HEL epitopes 25-43, 34-45, 112-124, and 116-129 was unchanged or even slightly diminished in RKKI cells. The presentation of the epitopes 25-43 and 34-45 was virtually insensitive to the lysosomotropic reagent chloroquine. Brefeldin A (BFA), which inhibits protein egress from the endoplasmic reticulum, blocked the presentation of all epitopes tested in RKKI cells. In contrast, in Ii-negative RKK cells only the presentation of the epitope HEL(46-61) was inhibited by BFA and the presentation of the epitopes 25-43 and 34-45 was only slightly impaired. These findings suggest that Ii may target class II molecules to selected endosomal subcompartments involved in the processing of different peptides derived from an endocytosed antigen. As a result, the enhancement of the class II-restricted presentation in Ii expressing cells appears to be epitope specific rather than antigen specific.

Sorting of MHC class II molecules and the associated invariant chain (Ii) in polarized MDCK cells

1997 ◽  
Vol 110 (5) ◽  
pp. 597-609 ◽  
Author(s):  
A. Simonsen ◽  
E. Stang ◽  
B. Bremnes ◽  
M. Roe ◽  
K. Prydz ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
Antigen Presentation ◽  
Mhc Class Ii ◽  
Mdck Cells ◽  
Intracellular Localization ◽  
Cytoplasmic Tail ◽  
Class Ii ◽  
Invariant Chain ◽  
Basolateral Targeting ◽  
Mhc Class Ii Molecules

Epithelial cells have been found to express MHC class II molecules in vivo and are able to perform class II-restricted antigen presentation. The precise intracellular localization of these molecules in epithelial cells has been a matter of debate. We have analyzed the polarized targeting of human MHC class II molecules and the associated invariant chain (Ii) in stably transfected MDCK cells. The class II molecules are located at the basolateral surface and in intracellular vesicles, both when expressed alone or together with Ii. Ii is located in basolateral endosomes and can internalize through the basolateral plasma membrane domain. We show that the cytoplasmic tail of Ii contains information for basolateral targeting as it is sufficient to redirect the apical protein neuraminidase (NA) to the basolateral surface. We find that the two leucine-based motifs (LI and ML) in the cytoplasmic tail of Ii are individually sufficient for endosomal sorting and basolateral targeting of Ii in MDCK cells. In addition, basolateral sorting information is located within the 10 membrane-proximal residues of the Ii cytoplasmic tail. As several different signals mediate basolateral sorting of the class II/Ii complex, a polarized distribution of these molecules may be an essential feature of antigen presentation in epithelial cells.

scholarly journals Antigen compartmentation and T helper cell tolerance induction.

1996 ◽  
Vol 183 (6) ◽  
pp. 2617-2626 ◽  
Author(s):  
S Oehen ◽  
L Feng ◽  
Y Xia ◽  
C D Surh ◽  
S M Hedrick
Keyword(s):  
Plasma Membrane ◽  
Mhc Class Ii ◽  
Antigen Processing ◽  
Tolerance Induction ◽  
Cell Receptor ◽  
Class Ii ◽  
Endocytic Pathway ◽  
Peptide Antigens ◽  
Processing Pathways ◽  
Bone Marrow Chimeras

The process of antigen recognition depends in part on the amount of peptide antigen available and the affinity of the T cell receptor for a particular peptide-major histocompatibility complex (MHC) molecule complex. The availability of self antigen is limited by antigen processing, which is compartmentalized such that peptide antigens presented by MHC class I molecules originate in the cytoplasm, whereas peptide antigens presented by MHC class II molecules are acquired from the endocytic pathway. This segregation of the antigen-processing pathways may limit the diversity of antigens that influence the development and selection of, e.g., CD4-positive, MHC class II-specific T cells. Selection in this case might involve only a subset of self-encoded proteins, specifically those that are plasma membrane bound or secreted. To study these aspects of immune development, we engineered pigeon cytochrome for expression in transgenic mice in two forms: one in which it was expressed as a type II plasma membrane protein, and a second in which it was targeted to the mitochondria after cytoplasmic synthesis. Experiments with these mice clearly show that tolerance is induced in the thymus, irrespective of antigen compartmentation. Using radiation bone marrow chimeras, we further show that cytoplasmic/mitochondrial antigen gains access to the MHC class II pathway by direct presentation. As a result of studying the anatomy of the thymus, we show that the amount of antigen and the affinity of the TCR affect the location and time point of thymocytes under-going apoptosis.

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