|
|
|
|
|
|
|
||
Articles |
Department of Pediatrics, National Jewish Center for Immunology, Denver, Colorado 80206
 |
Abstract |
|---|
 Top Abstract Materials and MethodsResults and DiscussionReferences |
|---|
subunit of the BCR, which further illustrates the unique nature of these endocytic vesicles.
The recognition of MHC class II–restricted antigens by antigen-specific T cells requires the proteolytic processing of protein antigens to immunogenic peptides by class II–positive antigen-presenting cells (1, 2). The first step in antigen processing by B cells involves B cell receptor (BCR)1–mediated internalization of antigen (3–5). BCR-internalized antigen is then proteolytically processed and the resultant peptides preferentially loaded onto newly synthesized class II molecules (6–8) from which the class II– associated invariant chain has been removed by the concerted action of acid proteases and the protein HLA-DM/ H-2M (9). The resultant peptide–class II complexes are then transported to the surface of the B cell.
The intracellular compartments where antigen processing occurs have only recently been characterized and there is considerable variation in the intracellular localization of class II molecules among different cell types. Many cells, such as human lymphoblasts and macrophages, sequester much of their class II in lysosomes or lysosome-like structures referred to as the MHC class II–enriched compartment (MIIC; reference 10). Although delivery of BCR-internalized antigen to MIIC has been demonstrated (11), the fate of the antigen delivered to these structures (i.e., complete degradation versus processing and binding to class II molecules) remains unknown.
In other professional antigen-presenting cells such as many murine B cell lines, there is little accumulation of class II in lysosomes under normal conditions (12–14). Instead, class II is found in endosomes and endosome-related structures, at least one population of which (class II vesicles [CIIV]) can be purified and physically separated from conventional endocytic and secretory organelles by cell fractionation techniques (14).
Although many or all endocytic, class II–containing vesicle populations may host the loading of peptides onto class II molecules, there may be important qualitative differences regarding the subcellular compartments where antigenic peptides are generated and efficiently loaded onto class II molecules. Specifically, although BCR-mediated antigen presentation appears to involve binding of peptide to newly synthesized class II molecules (6–8), presentation of fluid phase proteins by B cells appears to be able to occur via both newly synthesized and recycling class II molecules (7, 8, 15, 16), possibly reflecting differences in the intracellular sites of peptide generation and class II loading.
Additionally, not all receptors are equivalent at mediating antigen processing and presentation. In murine B cells, antigen internalized via the transferrin receptor (while presented more efficiently than soluble antigen) is presented 10–100 times less efficiently than the same antigen internalized via the BCR (17). This result may reflect the fact that the transferrin receptor has far more restricted access to intracellular class II compartments in B cells than does the BCR (11). Even more dramatic is the demonstration that a single amino acid substitution in the transmembrane region of the human IgM BCR (huBCR) can completely abolish the ability of this receptor to mediate efficient antigen processing and presentation without affecting BCR-mediated antigen endocytosis and bulk antigen degradation (18, 19). Thus, antigen uptake and degradation is necessary, but not sufficient, for antigen processing and presentation.
Thus, it has become important to determine the intracellular compartments to which physiologically important receptors (e.g., the BCR) deliver antigens. In this paper, we demonstrate that, within the time frame during which the intracellular events of BCR-mediated antigen processing are known to occur, BCR molecules and BCR-internalized antigen have access not only to predominantly class II– negative endosome and lysosomes, but also to a novel population of endocytic vesicles that are highly enriched in newly synthesized class II molecules (i.e., CIIV). Moreover, CIIV contain a putative marker protein, immunologically related to the Ig
Distribution of huBCR-internalized Antigen in A20µWT FFE Fractions.
Immuno-electron Microscopy Localization of huBCR-internalized Antigen and BCR Molecules to Isolated CIIV.
Steady-state Distribution of the PC-specific huBCR in A20µWT Cells.
Surface Labeling and Endocytosis.
Detection of Biotin-labeled huBCR Molecules in A20µWT FFE Fractions.
Steady-state Distribution of Ig
Immunological Relationship Between Ig subunit of the BCR, further illustrating the distinct nature of these endocytic vesicles.
Materials and Methods
Top
Abstract
Materials and Methods
Results and Discussion
References
Cell Culture and Fractionation.
A20µWT (i.e., A20 cells expressing a transfected, phosphorylcholine (PC)–specific human mIgM BCR (huBCR); reference 19) were cultured in MEM, 5% FBS, 50 µM 2-mercaptoethanol, and 500 µg/ml G418. A20µWT cells were homogenized, fractionated by free flow electrophoresis (FFE), and the distribution of plasma membrane, lysosomes, and CIIV was determined as previously reported (14).
A20µWT cells (2 x 108 total cells) were incubated at 4 x 107 cells/ml for 30 min at 37°C in media containing 2 µg/ml PC-modified Fab fragments of rabbit 
globulin labeled with 125I (PC–RGG–125I [2µCi 125I/µg PC–RGG]), homogenized, fractionated by FFE, and the distribution of the plasma membrane and lysosomes was determined. The distribution of PC–RGG–125I was determined by counting each FFE fraction in a counter. Background counts (<100 cpm) were subtracted from the counts for each sample and the results normalized to a maximum value of 1.00.
A20µWT cells were incubated in 400-nM PC-modified ovalbumin (PC–OVA) for 20 min at 37°C, homogenized, and fractionated by FFE. Isolated CIIV, as well as endosomes–lysosomes, were processed for immuno-electron microscopy (immunoEM) as previously described (14). Cryosections were stained with rabbit IgG specific for either murine class II (14), murine IgG (315-005-046; Jackson Immunologicals, West Grove, PA), human IgM (309-005-095; Jackson Immunologicals; 309-005-095), or ovalbumin (RaOVALBUMI; East Acres Biologicals, Southbridge, MA). Antibody to class II molecules, anti-ovalbumin, and both anti-Ig antibodies were visualized with either 1, 5, or 10 nm protein A–gold (14).
A20µWT cells were homogenized and fractionated by FFE. 200 µl of each FFE fraction, along with 50 µl of homogenization buffer containing 5% Triton X-100 and 0.5 mg/ml BSA, was added to a PC–BSA-coated 96-well plate and the samples allowed to bind for 6 h at 4°C. The plates were washed and probed with rabbit anti–human IgM (1:1,000; 309-005-095; Jackson Immunologicals) followed by horseradish peroxidase (HRP)–labeled goat anti–rabbit IgG (1:1,000; 31462; Pierce Chemical Co., Rockford, IL). Bound goat anti–rabbit Ig–HRP was detected by addition of 200 µl of 0.5 mg/ml O-phenylenediamine and 0.015% H2O2 in borate buffer. After sufficient time, 50 µl of 1N HCl was added and the absorbance (OD 490 nm) measured. The absorbance above background is reported (background = 
0.200 OD 490 nm).
A20µWT cells were collected by centrifugation and washed two times with PBS. Cells were labeled for 15 min at 108 viable cells/ml in PBS pH 7.5 containing 1 mg/ml sulfosuccinimidyl-6-(biotinamido) hexanoate (NHS– LC–biotin; Pierce Chemical Company; 21335). The labeling was quenched by addition of 5–10 vol of 10 mM lysine in PBS. The labeled cells were pelleted and then washed twice in PBS 0.1% BSA before incubation at 37°C in complete media containing 1 µM PC–OVA (Cells were >98% viable after labeling, washing, and incubation).
FFE fractions (200 µl) from biotin-labeled/PC–OVA pulsed A20µWT cells were added to NeutrAvidin (50 µg/ml; 31000; Pierce)-coated plates along with 50 µl of homogenization buffer containing 5% Triton X-100 and 0.5 mg/ml BSA and allowed to bind for 6 h at 4°C. The plate was washed and probed with rabbit anti–human IgM (1:1,000, 309-005-095; Jackson Immunologicals) followed by HRP-labeled goat anti–rabbit Ig (1:1,000; 31462; Pierce). Bound goat anti–rabbit Ig–HRP was detected as described above. The absorbance above background is reported (background = 0.100 OD 490 nm). in A20µWT Cells.
Individual A20µWT FFE fractions were concentrated by centrifugation and analyzed by SDS-PAGE and Western blotting (14) with a rabbit antiserum raised against intact, full length, bacterially expressed murine Ig (rabbit anti-Ig, 1:5,000). Binding of rabbit anti-Ig was detected with HRP-labeled goat anti–rabbit Ig (1:5,000; 31462; Pierce) and enhanced chemiluminescence (ECL; 34080; Pierce). and p50Ig.
A20µWT low density membranes (LDM) were separated by preparative SDS-PAGE and electroblotted onto nitrocellulose. The blot was probed with rabbit anti-Ig (1:1,000) and then extensively washed. The regions of the blot containing either Iga and p50Ig were individually excised and bound antibodies eluted with 500 µl of 0.1 M glycine, pH 2.5. The eluted antibody was neutralized with 100 µl, 1.0 M Tris, pH 8.0, and 4.5 ml of blotting buffer. The affinity-purified antibody was used to probe Western blots of total A20µWT LDM and binding revealed with HRP-labeled goat anti–rabbit Ig (1:5,000; 31462; Pierce) and ECL (34080; Pierce).
Results and Discussion
Top
Abstract
Materials and Methods
Results and Discussion
References
BCR-mediated Delivery of Antigen to Endosome and CIIV.
The murine A20µWT cell line (19) was used as a model cell for these studies. A20µWT cells express a PC-specific huBCR as well as an endogenous murine IgG2a BCR (muBCR) and the ability of these cells to process and present antigens via both BCRs has been well characterized (4, 19). Importantly, A20µWT cells localize only a small fraction (<10%) of their total class II to intracellular membranes (13, 14), with little class II present in high density, hydrolase-rich lysosomes (14). Owing to the lack of class II molecules in the lysosomes of these cells, it is easy to distinguish, by FFE, a distinctive population of low density, novel endocytic vesicles (i.e., CIIV) that are enriched in newly synthesized class II molecules (14). Moreover, comparative analysis of A20 (14, 23) and A20µWT cells (Figs. 1, 2 A, and 4; data not shown) demonstrates that CIIV isolated from both cells possesses the same morphological, biophysical, biochemical, and immunological characteristics (e.g., class II–positive, lgp110-negative, β-Hexosaminidase– negative multivesicular membrane structures with a characteristic electrophoretic mobility). Importantly, in A20µWT cells, BCR-mediated antigen processing and peptide loading of class II molecules can occur exclusively in low density endocytic structures without the involvement of high density lysosomal structures (13, 14).
|
|
|
This biochemical analysis, along with our previous observations that CIIV-containing FFE fractions consist almost entirely of class II–positive vesicles, strongly suggested that BCR-internalized antigen can gain access to CIIV. To demonstrate this point directly, and rule out the possibility that the BCR-internalized antigen present in the CIIV-containing FFE fractions was contained exclusively in class II–negative, β-Hexosaminidase–positive lysosomes, CIIV-containing FFE fractions from antigen (i.e., PC–OVA)– pulsed A20µWT cells were examined by multiple label immunoEM (14). As shown in Fig. 2 A, huBCR-internalized antigen (arrow) was present in CIIV isolated from antigen-pulsed B cells. Additionally, immunoEM analysis of endosome/lysosome–enriched FFE fractions demonstrated that BCR-internalized antigen was present both in class II–negative endosomes and lysosomes as well as class II–positive endosomes (Fig. 2, B and C, respectively). Quantitation of these immunoEM samples (Table 1) revealed that the vast majority (71–85%) of CIIV were endocytic (i.e., accessible by BCR-internalized antigen) and that the bulk (58–90%) of the endocytic vesicles within the CIIV-containing FFE fractions were class II positive. On the contrary, only 34% of the endocytic vesicles in the endosome/lysosome–containing FFE fractions were class II–positive endosomes, with the majority of the antigen-containing vesicles being class II–negative endosomes and lysosomes.
|
|
Because we have previously demonstrated that newly synthesized BCR molecules do not traffic through CIIV before arrival at the cell surface (14), these results strongly suggest that the BCR molecules found in CIIV are derived from the PM by endocytosis and suggest that antigen is delivered to this compartment while bound to these internalized BCR molecules. To demonstrate directly that cell surface BCR molecules, internalized in the presence of polyvalent antigen, are delivered to CIIV, A20µWT were surface labeled with biotin, incubated for various times at 37°C in the presence of polyvalent antigen (i.e., PC–OVA), homogenized, and then fractionated by FFE. The level of biotin-labeled (i.e., internalized) huBCR molecules in each FFE fraction was then determined by a human IgM-specific, avidin-capture ELISA. As shown in Fig. 3 B, after 20 min of incubation, internalized huBCR molecules could be detected in CIIV-containing FFE fractions (fractions 53–60) as well as those enriched in endosomes and lysosomes (fractions 45–52). Surprisingly, even though we had found that, under similar conditions, a vast majority of the BCR-internalized antigen was ultimately delivered to endosomes and lysosomes (see Fig. 1), a significant fraction of the internalized huBCR molecules were found in CIIV-containing FFE fractions. Therefore, we suggest that a portion, and possibly all, of the huBCR molecules detected in CIIV by immunoEM (Fig. 4) were derived from the PM after endocytosis, further supporting the contention that antigen is delivered to CIIV while bound to the BCR. Interestingly, as suggested by the presence of huBCR molecules in CIIV isolated from nonantigen-pulsed A20µWT cells, preliminary analysis of the constitutive endocytosis and trafficking of the huBCR of A20µWT cells suggests that BCR endocytosis and delivery to CIIV can occur in the absence of antigen cross-linking (Drake, J.R., unpublished results).
Subcellular Distribution of BCR Subunits and Identification of a Putative CIIV Marker Protein Immunologically Related to Ig.
Although both BCR molecules and BCR-internalized antigen clearly gained access to CIIV, as well as endosomes and class II–negative lysosomes, the extent to which these molecules are, or are not, selectively targeted to CIIV remains unclear. To begin to address whether there is any selective targeting of BCR molecules or antigen–BCR complexes to CIIV, we first attempted to determine whether there is any difference in the subunit composition of the BCR molecules present in PM, endosome and lysosome, or CIIV-containing FFE fractions.
To this end, we examined the steady state distribution of the Ig subunit of the BCR. As shown in Fig. 5 A, the majority of the 32-kD Ig protein (arrow B) is present in PM-containing FFE fractions with lesser amounts detected in endosome–lysosome and CIIV-containing fractions. The same distribution was also found for the Igβ subunit of the BCR as well as the heavy and light chain subunits of both the huBCR and muBCR (data not shown), suggesting that the subunit composition of the BCRs in these compartments is similar. Surprisingly, the anti-Ig antiserum also recognized a second, 50-kD protein (Fig. 5 A, arrow A) that appears to be selectively enriched in CIIV-containing FFE fractions.
|
(as opposed to being recognized by antibodies of a second specificity present in the rabbit anti-Ig antiserum), we affinity-purified antibodies to both Ig and the 50-kD protein on Western blots of A20µWT LDM, and tested the specificity of these purified antibodies. As shown in Fig. 5 B, the unfractionated rabbit anti-Ig antiserum recognized both Ig as well as the 50-kD protein present in unfractionated A20µWT LDM. Although affinity-purified anti-Ig antibody failed to recognize the 50-kD protein (occasionally, reactivity of the affinity-purified anti-Ig toward the 50-kD protein was observed although the results were variable, possibly owing to removal of low affinity/highly cross-reactive antibodies by the affinity purification protocol), affinity-purified antibody against the 50-kD protein recognized both Ig as well as the 50-kD protein. Because this antiserum was originally generated against recombinant whole murine Ig, these results demonstrate that at least some anti-Ig antibodies specifically recognize the 50-kD putative CIIV-marker protein, demonstrating that these proteins are immunologically related (i.e., that Ig and the 50-kD protein minimally share one cross-reactive epitope). Additionally, an antiserum raised against the cytoplasmic tail of Ig (20) also demonstrated cross-reactivity to the 50-kD protein, suggesting an immunological relationship between the cytoplasmic tail of Ig and some region of the 50-kD protein (data not shown).
Because the 50-kD putative CIIV-marker protein does not exhibit a decrease in apparent molecular weight upon treatment with either endoglycosidase H or F (Drake, J.R., unpublished results), we have foregone the more traditional gp50 designation in favor of p50Ig (the Ig superscript indicates the immunological relationship of the protein to the Ig subunit of the BCR). Although the structure and function of p50Ig remains unknown, it is unlikely to represent a highly modified form of Ig, since it was also detected in Ig-negative J774 macrophage-like cell line (Drake, J.R., unpublished results). Moreover, p50Ig is unlikely to be an artifact of proteolytic activity because it can be detected in detergent extracts of whole cells prepared in the presence of a cocktail of protease inhibitors (Drake, J.R., unpublished results). Additionally, preliminary analysis of J774 cells and the murine B cell hybridoma 2C3E1 (21) suggests a restricted distribution of p50Ig to CIIV in these cells (Drake, J.R., and P. Webster, unpublished results).
Considering the presence of p50Ig in the BCR-negative J774 macrophage-like cell line, p50Ig, unlike Ig, may not be a component of the BCR protein complex. Correspondingly, preliminary analysis has failed to reveal any physical association between p50Ig and the BCR of A20µWT cells (Drake, J.R., unpublished results). Therefore, a more thorough understanding of the possible function of p50Ig will have to await its eventual purification and sequencing or cDNA cloning. Interestingly, previous Southern blot analysis of the murine MB-1 gene (which codes for Ig) indicated the presence of an additional, Ig-related gene (22), possibly that coding for p50Ig. Most importantly, the identification of p50Ig as a putative marker for CIIV graphically demonstrates the unique biochemical nature of these novel class II-containing vesicles and provides us with a tool to study their origin, fate, and relationship to other intracellular compartments.
Previously, we demonstrated that in murine B cells, class II molecules are restricted to relatively early endocytic compartments (i.e., endosomes and CIIV), with little or no class II found in high density lysosomes (14). Because peptide–class II complexes have been demonstrated to form only in low density (i.e., nonlysosomal) compartments in these cells (13, 14), the presence of BCR-internalized antigen in both class II–positive endosomes as well as CIIV suggests that antigen processing and class II peptide loading may occur at either or both of these sites. Given the predominant role of newly synthesized class II molecules in BCR-mediated antigen processing and presentation (6–8) and the fact that CIIV are an intermediate in the pathway of transport of newly synthesized class II molecules to the cell surface (14, 23), our results strongly suggest a role for CIIV in BCR-mediated antigen processing and class II peptide loading.
Why might B cells possess a novel endocytic compartment for the processing of BCR-internalized antigens? In contrast with endosomes, lysosomes, and MIIC, which readily accumulate nonselectively internalized fluid phase endocytic tracers (10, 24), CIIV do not readily accumulate proteins internalized by fluid phase endocytosis (14). Within endosomes, lysosomes, and MIIC, the vast excess of nonantigenic peptides (i.e., peptides derived from the proteolytic degradation of fluid phase plasma proteins) may effectively compete with antigenic peptides (i.e., peptides derived from BCR-internalized antigen) for binding to class II molecules, preventing the efficient formation of antigenic peptide–class II complexes within these compartments. The absence of these nonantigenic peptides from CIIV may allow for more efficient formation of antigenic peptide–class II complexes in these vesicles. Additionally, because peptide loading onto class II is a relatively slow process (25), the relatively slow transport of newly synthesized class II molecules through CIIV, with class II molecules residing in CIIV for up to 2 h (14), may provide the necessary time for the loading of these class II molecules with antigen-derived peptides.
|
Acknowledgments |
|---|
Submitted: 11 June 1997
Revised: 19 August 1997
1 Abbreviations used in this paper: BCR, B cell receptor; CIIV, class II vesicles; ECL, enhanced chemiluminescence; FFE, free flow electrophoresis; huBCR, phosphorylcholine-specific human mIgM BCR; immunoEM; immuno-electron microscopy; LDM, low density membranes; MIIC, MHC class II–enriched compartment; muBCR, murine IgG2a BCR; NHS–LC–biotin, sulfosuccinimidyl-6-(biotinamido) hexanoate; PC, phosphorylcholine; PC–RGG–125I, PC-modified Fab fragments of rabbit
globulin labeled with 125I; PC–OVA, PC-modified ovalbumin; PM, plasma membrane. 
|
References |
|---|
|
TopAbstractMaterials and MethodsResults and DiscussionReferences |
|---|
1 Wolf PR & Ploegh HL. How MHC class II molecules acquire peptide cargo: biosynthesis and trafficking through the endocytic pathway, Annu Rev Cell Dev Biol, 1995, 11, 267–306.[Medline]
2 Germain RN & Margulies DH. The biochemistry and cell biology of antigen processing and presentation, Annu Rev Immunol, 1993, 11, 403–450.[Medline]
3 Grey HM, Colon SM & Chesnut RW. Requirements for the processing of antigen by antigen-presenting B cells. II. Biochemical comparison of the fate of antigen in B cell tumors and macrophages, J Immunol, 1982, 129, 2389–2395.[Medline]
4 Chesnut RW, Colon SM & Grey HM. Requirements for the processing of antigens by antigen-presenting B cells. I. Functional comparison of B cell tumors and macrophages, J Immunol, 1982, 129, 2382–2388.[Medline]
5 Lanzavecchia A. Antigen-specific interaction between T and B cells, Nature (Lond), 1985, 314, 537–539.[Medline]
6 Davidson HW, Reid PA, Lanzavecchia A & Watts C. Processed antigen binds to newly synthesized MHC class II molecules in antigen-specific B lymphocytes, Cell, 1991, 67, 105–116.[Medline]
7 Kakiuchi T, Watanabe M, Hozumi N & Nariuchi H. Differential sensitivity of specific and nonspecific antigen-presentation by B cells to a protein synthesis inhibitor, J Immunol, 1990, 145, 1653–1658.[Abstract]
8 Kakiuchi T, Takatsuki A, Watanabe M & Nariuchi H. Inhibition by brefeldin A of the specific B cell antigen presentation to MHC class II–restricted T cells, J Immunol, 1991, 147, 3289–3295.[Abstract]
9 Roche PA. HLA-DM: an in vivo facilitator of MHC class II peptide loading, Immunity, 1995, 3, 259–262.[Medline]
10 Peters PJ, Neefjes JJ, Oorschot V, Ploegh HL & Geuze HJ. Segregation of MHC class II molecules from MHC class I molecules in the Golgi complex for transport to lysosomal compartments, Nature (Lond), 1991, 349, 669–676.[Medline]
11 West MA, Lucocq JM & Watts C. Antigen processing and class II MHC peptide-loading compartments in human B-lymphoblastoid cells, Nature (Lond), 1994, 369, 147–151.[Medline]
12 Brachet V, Raposo G, Amigorena S & Mellman I. Ii chain controls the transport of major histocompatability complex class II molecules to and from lysosomes, J Cell Biol, 1997, 137, 51–65.
13 Barnes KA & Mitchell RN. Detection of functional class II–associated antigen: role of a low density endosomal compartment in antigen processing, J Exp Med, 1995, 181, 1715–1727.
14 Amigorena S, Drake JR, Webster P & Mellman I. Transient accumulation of new class II MHC molecules in a novel endocytic compartment in B lymphocytes, Nature (Lond), 1994, 369, 113–120.[Medline]
15 Soreng KM, Moore JC, Sherman MA & Jensen PE. Requirement for protein synthesis in antigen processing by B cells, Cell Immunol, 1994, 157, 277–290.[Medline]
16 St. Pierre Y & Watts TH. MHC class II–restricted presentation of native protein antigen by B cells is inhibitable by cycloheximide and brefeldin A, J Immunol, 1990, 145, 812–818.[Abstract]
17 Niebling WL & Pierce SK. Antigen entry into early endosomes is insufficient for MHC class II processing, J Immunol, 1993, 150, 2687–2697.[Abstract]
18 Mitchell RN, Barnes KA, Grupp SA, Sanchez M, Misulovin Z, Nussenzweig MC & Abbas AK. Intracellular targeting of antigens internalized by membrane immunoglobulin in B lymphocytes, J Exp Med, 1995, 181, 1705–1714.
19 Shaw AC, Mitchell RN, Weaver YK, Campos TJ, Abbas AK & Leder P. Mutations of immunoglobulin transmembrane and cytoplasmic domains: effects on intracellular signaling and antigen presentation, Cell, 1990, 63, 381–392.[Medline]
20 Clark MR, Campbell KS, Kazlauskas A, Johnson SA, Hertz M, Potter TA, Pleiman C & Cambier JC. The B cell antigen receptor complex: association of Ig-alpha and Ig-beta with distinct cytoplasmic effectors, Science (Wash DC), 1992, 258, 123–126.
21 Drake JR, Repasky EA & Bankert RB. Endocytosis of antigen, anti-idiotype, and anti-immunoglobulin antibodies and receptor re-expression by murine B cells, J Immunol, 1989, 143, 1768–1776.[Abstract]
22 Kashiwamura S, Koyama T, Matsuo T, Steinmetz M, Kimoto M & Sakaguchi N. Structure of the murine mb-1 gene encoding a putative sIgM-associated molecule, J Immunol, 1990, 145, 337–343.[Abstract]
23 Amigorena S, Webster P, Drake J, Newcomb J, Cresswell P & Mellman I. Invariant chain cleavage and peptide loading in major histocompatibility complex class II vesicles, J Exp Med, 1995, 181, 1729–1741.
24 Peters PJ, Raposo G, Neefjes JJ, Oorschot V, Leijendekker RL, Geuze HJ & Ploegh HL. Major histocompatibility complex class II compartments in human B lymphoblastoid cells are distinct from early endosomes, J Exp Med, 1995, 182, 325–334.
25 Jensen PE. Enhanced binding of peptide antigen to purified class II major histocompatibility glycoproteins at acidic pH , J Exp Med, 1991, 174, 1111–1120.
26 Shaw MK, Tilney LG & Musoke AJ. The entry of Theileria parva sporozoites into bovine lymphocytes: evidence for MHC class I involvement, J Cell Biol, 1991, 113, 87–101.
CiteULike 
Complore 
Connotea 
Del.icio.us 
Digg 
Facebook 
Reddit 
Technorati 
Twitter What's this?
This article has been cited by other articles:
| |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
| TABLE OF CONTENTS |
|
