scholarly journals Rescue of Daudi cell HLA expression by transfection of the mouse beta 2-microglobulin gene.

1988 ◽  
Vol 167 (2) ◽  
pp. 288-299 ◽  
Author(s):  
R H Seong ◽  
C A Clayberger ◽  
A M Krensky ◽  
J R Parnes
Keyword(s):  
Cell Surface ◽  
Surface Expression ◽  
Class I ◽  
Cell Surface Expression ◽  
Daudi Cell ◽  
Heavy Chains ◽  
Daudi Cells ◽  
Beta 2 ◽  
Beta 2 Microglobulin ◽  
Hla Molecules

The Daudi cell line is a B-lymphoblastoid line derived from a Burkitt lymphoma. Daudi cells lack cell surface expression of class I HLA molecules despite the presence of intracellular class I heavy chains. They have a defect in the gene encoding beta 2-microglobulin (beta 2m), resulting in lack of translatable mRNA for this protein. It has been thought that this deficiency is responsible for the lack of cell surface class I expression. However, data have recently been presented demonstrating that at least one mouse class I heavy chain can be expressed on the cell surface in the absence of beta 2m. These results raised the questions of whether the lack of beta 2m is the only defect in Daudi and whether transfer of this single gene could restore surface class I expression. We found that transfection of the mouse beta 2m gene into Daudi indeed rescued cell surface expression of class I HLA molecules, and that these molecules could be recognized both by monomorphic and allospecific mAbs. CTL clones specific for HLA-B17 or a determinant shared by HLA-B17 and HLA-A2 killed the Daudi cells transfected with the beta 2m gene, but not untransfected Daudi or Daudi transfected with vector alone. Mouse beta 2m on the transfected Daudi cells could exchange with human beta 2m when the cells were incubated in human serum. This exchange did not alter the ability of the cells to be killed by the specific CTLs. These results demonstrate that the lack of beta 2m is the sole reason for lack of surface class I molecules in Daudi cells, and that beta 2m is required for cell surface expression of the specific class I heavy chains of Daudi.

scholarly journals Increase in positive selection of CD8+ T cells in TAP1-mutant mice by human beta 2-microglobulin transgene.

1995 ◽  
Vol 181 (2) ◽  
pp. 787-792 ◽  
Author(s):  
H Martien van Santen ◽  
A Woolsey ◽  
P G Rickardt ◽  
L Van Kaer ◽  
E J Baas ◽  
...  
Keyword(s):  
T Cells ◽  
Cell Surface ◽  
Mhc Class I ◽  
Cd8 T Cells ◽  
Surface Expression ◽  
Class I ◽  
Cell Surface Expression ◽  
Beta 2 ◽  
Beta 2 Microglobulin ◽  
Selection Of

Mice harboring a deletion of the gene encoding the transporter associated with antigen presentation-1 (TAP1) are impaired in providing major histocompatibility complex (MHC) class I molecules with peptides of cytosolic origin and lack stable MHC class I cell surface expression. They consequently have a strongly reduced number of CD8+ T cells. To examine whether selection of CD8+ T cells is dependent on TAP-dependent peptides, we partially restored MHC class I cell surface expression in TAP1-deficient mice by introduction of human beta 2-microglobulin. We show that selection of functional CD8+ T cells can be augmented in vivo in the absence of TAP1-dependent peptides.

scholarly journals Abrogation of cell surface expression of human class I transplantation antigens by an adenovirus protein in Xenopus laevis oocytes.

1985 ◽  
Vol 101 (2) ◽  
pp. 540-547 ◽  
Author(s):  
L Severinsson ◽  
P A Peterson
Keyword(s):  
Cell Surface ◽  
Heavy Chain ◽  
Viral Protein ◽  
Surface Expression ◽  
Class I ◽  
Cell Surface Expression ◽  
Xenopus Laevis Oocytes ◽  
Human Class ◽  
Heavy Chains ◽  
Transplantation Antigens

Class I transplantation antigens form complexes with a virus protein encoded in the early region E3 of the adenovirus-2 genome. The interaction between this viral glycoprotein, E19, and nascent human class I antigens has been examined by microinjecting purified mRNA into Xenopus laevis oocytes. Both E19 and the two class I antigen subunits, the heavy chain and beta 2-microglobulin (beta 2M), were efficiently translated. The heavy chains did not become terminally glycosylated, as monitored by endoglycosidase H digestion, and were not expressed on the oocyte surface unless they were associated with beta 2M. The E19 protein did not become terminally glycosylated, and we failed to detect this viral protein on the surface of the oocytes. Co-translation of heavy chain and E19 mRNA demonstrated that the two proteins associate intracellularly. However, neither protein appeared to be transported to the trans-Golgi compartment. Similar observations were made in adenovirus-infected HeLa cells. Heavy chains bound to beta 2M became terminally glycosylated in oocytes in the presence of low concentrations of E19. At high concentrations of the viral protein, no carbohydrate modifications and no cell surface expression of class I antigens were apparent. Thus, beta 2M and E19 have opposite effects on the intracellular transport of the heavy chains. These data suggest that adenovirus-2 may impede the cell surface expression of class I antigens to escape immune surveillance.

scholarly journals Amino acid sequences in the alpha 1 domain and not glycosylation are important in HLA-A2/beta 2-microglobulin association and cell surface expression.

1987 ◽  
Vol 7 (3) ◽  
pp. 982-990 ◽  
Author(s):  
J Santos-Aguado ◽  
P A Biro ◽  
U Fuhrmann ◽  
J L Strominger ◽  
J A Barbosa
Keyword(s):  
Amino Acid ◽  
Critical Role ◽  
Surface Expression ◽  
Amino Acid Sequences ◽  
Cell Surface Expression ◽  
Amino Acid Substitutions ◽  
Heavy Chains ◽  
Beta 2 ◽  
Beta 2 Microglobulin ◽  
Asparagine Residue

The role of the single carbohydrate moiety present on the HLA-A2 molecule was studied by introducing several amino acid substitutions (by site-directed mutagenesis of the HLA-A2 gene) in the consensus glycosylation sequence Asn-X-Ser. Two different amino acid substitutions of the asparagine residue at position 86 (glutamine and aspartic acid) resulted in the synthesis of ca. 39,000-molecular-weight nonglycosylated heavy chains that were detected in the cytoplasm but not on the surface of mouse L-cell transfectants. However, a low level of surface expression was detected following transfection of human (rhabdomyosarcoma) cells or mouse L cells containing human beta 2-microglobulin. The defect in surface expression was not due to the absence of the glycan moiety, since the substitution of a glycine for a serine at amino acid 88 did not have the same drastic effect in the presence of human beta 2-microglobulin. These and other data suggest that the asparagine residue may play a critical role in the conformation of the HLA heavy chain and its interaction with beta 2-microglobulin. Immunofluorescence microscopy following permeabilization of the transfectants demonstrated that the unglycosylated HLA heavy chains are sequestered in an unidentified cellular compartment that is different from the Golgi structure.

scholarly journals Impaired intracellular transport and cell surface expression of nonpolymorphic HLA-E: evidence for inefficient peptide binding.

1992 ◽  
Vol 176 (4) ◽  
pp. 1083-1090 ◽  
Author(s):  
M Ulbrecht ◽  
J Kellermann ◽  
J P Johnson ◽  
E H Weiss
Keyword(s):  
Cell Surface ◽  
Intracellular Transport ◽  
Hla Class I ◽  
Surface Expression ◽  
Myeloma Cell ◽  
Peptide Ligands ◽  
Class I ◽  
Cell Surface Expression ◽  
Trimolecular Complex ◽  
Beta 2 Microglobulin

The assembly of the classical, polymorphic major histocompatibility complex class I molecules in the endoplasmic reticulum requires the presence of peptide ligands and beta 2-microglobulin (beta 2m). Formation of this trimolecular complex is a prerequisite for efficient transport to the cell surface, where presented peptides are scanned by T lymphocytes. The function of the other class I molecules is in dispute. The human, nonclassical class I gene, HLA-E, was found to be ubiquitously transcribed, whereas cell surface expression was difficult to detect upon transfection. Pulse chase experiments revealed that the HLA-E heavy chain in transfectants, obtained with the murine myeloma cell line P3X63-Ag8.653 (X63), displays a significant reduction in oligosaccharide maturation and intracellular transport compared with HLA-B27 in corresponding transfectants. The accordingly low HLA-E cell surface expression could be significantly enhanced by either reducing the culture temperature or by supplementing the medium with human beta 2m, suggesting inefficient binding of endogenous peptides to HLA-E. To analyze whether HLA-E binds peptides and to identify the corresponding ligands, fractions of acid-extracted material from HLA-E/X63 transfectants were separated by reverse phase HPLC and were tested for their ability to enhance HLA-E cell surface expression. Two fractions specifically increased the HLA class I expression on the HLA-E transfectant clone.

scholarly journals Cell surface expression of beta 2-microglobulin (beta 2m) correlates with stages of differentiation in B cell tumours.

1987 ◽  
Vol 40 (5) ◽  
pp. 486-489 ◽  
Author(s):  
R A Jones ◽  
C S Scott ◽  
D R Norfolk ◽  
A N Stark ◽  
J A Child
Keyword(s):  
Cell Surface ◽  
B Cell ◽  
Surface Expression ◽  
Cell Surface Expression ◽  
Beta 2 ◽  
Beta 2 Microglobulin

Amino acid sequences in the alpha 1 domain and not glycosylation are important in HLA-A2/beta 2-microglobulin association and cell surface expression

1987 ◽  
Vol 7 (3) ◽  
pp. 982-990
Author(s):  
J Santos-Aguado ◽  
P A Biro ◽  
U Fuhrmann ◽  
J L Strominger ◽  
J A Barbosa
Keyword(s):  
Amino Acid ◽  
Critical Role ◽  
Surface Expression ◽  
Amino Acid Sequences ◽  
Cell Surface Expression ◽  
Amino Acid Substitutions ◽  
Heavy Chains ◽  
Beta 2 ◽  
Beta 2 Microglobulin ◽  
Asparagine Residue

The role of the single carbohydrate moiety present on the HLA-A2 molecule was studied by introducing several amino acid substitutions (by site-directed mutagenesis of the HLA-A2 gene) in the consensus glycosylation sequence Asn-X-Ser. Two different amino acid substitutions of the asparagine residue at position 86 (glutamine and aspartic acid) resulted in the synthesis of ca. 39,000-molecular-weight nonglycosylated heavy chains that were detected in the cytoplasm but not on the surface of mouse L-cell transfectants. However, a low level of surface expression was detected following transfection of human (rhabdomyosarcoma) cells or mouse L cells containing human beta 2-microglobulin. The defect in surface expression was not due to the absence of the glycan moiety, since the substitution of a glycine for a serine at amino acid 88 did not have the same drastic effect in the presence of human beta 2-microglobulin. These and other data suggest that the asparagine residue may play a critical role in the conformation of the HLA heavy chain and its interaction with beta 2-microglobulin. Immunofluorescence microscopy following permeabilization of the transfectants demonstrated that the unglycosylated HLA heavy chains are sequestered in an unidentified cellular compartment that is different from the Golgi structure.

scholarly journals Variations in MHC class I antigen presentation and immunopeptidome selection pathways

F1000Research ◽  
2020 ◽  
Vol 9 ◽  
pp. 1177
Author(s):  
Anita J. Zaitouna ◽  
Amanpreet Kaur ◽  
Malini Raghavan
Keyword(s):  
Cell Surface ◽  
Antigen Presentation ◽  
Surface Expression ◽  
Structural Features ◽  
Class I ◽  
Cell Surface Expression ◽  
Immune Recognition ◽  
Antigen Receptors ◽  
Class I Mhc ◽  
Mhc I

Major histocompatibility class I (MHC-I) proteins mediate immunosurveillance against pathogens and cancers by presenting antigenic or mutated peptides to antigen receptors of CD8+ T cells and by engaging receptors of natural killer (NK) cells. In humans, MHC-I molecules are highly polymorphic. MHC-I variations permit the display of thousands of distinct peptides at the cell surface. Recent mass spectrometric studies have revealed unique and shared characteristics of the peptidomes of individual MHC-I variants. The cell surface expression of MHC-I–peptide complexes requires the functions of many intracellular assembly factors, including the transporter associated with antigen presentation (TAP), tapasin, calreticulin, ERp57, TAP-binding protein related (TAPBPR), endoplasmic reticulum aminopeptidases (ERAPs), and the proteasomes. Recent studies provide important insights into the structural features of these factors that govern MHC-I assembly as well as the mechanisms underlying peptide exchange. Conformational sensing of MHC-I molecules mediates the quality control of intracellular MHC-I assembly and contributes to immune recognition by CD8 at the cell surface. Recent studies also show that several MHC-I variants can follow unconventional assembly routes to the cell surface, conferring selective immune advantages that can be exploited for immunotherapy.

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