共查询到7条相似文献,搜索用时 0 毫秒
1.
Recent studies have shown enhanced antitumor efficacy with adenoviruses that either lack E1B-19K or overexpress E3-11.6K (also known as adenoviral death protein). E1B-19K is a well-characterized anti-apoptotic protein, and viruses with E1B-19K deletions show increased cytopathicity. However, the mechanism of cell killing by E3-11.6K, which plays an important role in killing infected cells and virion release, is not well characterized. To understand the mechanism of cell killing following E3-11.6K overexpression, we constructed a recombinant adenovirus, Ad-ME, by introducing viral major late promoter upstream of the E3-11.6K sequence. Similar to the E1B-19K-deleted virus, E1B/19K-, Ad-ME induced cell death to a greater extent than the wild-type virus. Cell shrinkage, membrane blebbing, activation of caspases 3 and 9, cleavage of poly(ADP-ribose)polymerase (PARP), DNA degradation, and ratio of ADP to ATP in Ad-ME-infected cells indicated that apoptosis contributes to cell death following E3-11.6K overexpression. However, the levels of activation of caspases 3 and 9 were lower in cells infected with Ad-ME compared to those infected with E1B/19K-. Furthermore, cell killing by Ad-ME was not effectively inhibited by Z-VAD-FMK, a general caspase inhibitor. Taken together, our results suggest both caspase-dependent and caspase-independent mechanisms of cell killing due to overexpression of E3-11.6K. 相似文献
2.
Joyce C. Solheim Nancy A. Johnson Beatriz M. Carreno Wen-Rong Lie Ted H. Hansen 《European journal of immunology》1995,25(11):3011-3016
With β2-microglobulin? (β2m?) cell lines such as R1E/Db, the surface expression of class I major histocompatibility complex molecules is greatly impaired, and class I molecules that are on the surface are generally misfolded. To determine whether β2m must be continually present with the class I heavy chain for the class I molecule to reach the surface in a folded conformation, a sequence encoding an endoplasmic reticulum (ER) retention signal (KDEL) was attached onto the 3′ end of a β2m cDNA. After this chimeric cDNA was transfected into R1E/Db cells, β2m-KDEL protein was detectable by an anti-β2m serum within the cells but not at the cell surface. Interestingly, R1E/Db cells transfected with β2m-KDEL were found to express a high level of conformationally correct Db molecules at the cell surface. This observation implies that β2m has a critical and temporal role in the de novo folding of the class I heavy chain. We propose that the critical time for β2m association is when the class I molecule is docked with the transporter associated with antigen processing (TAP) and first interacts with peptide. 相似文献
3.
L. J. Tan Stephanie Ceman Alexander Chervonsky Juan Rodriguez-Paris Theodore L. Steck Andrea J. Sant 《European journal of immunology》1997,27(6):1479-1488
The molecular mechanisms that regulate sorting of major histocompatibility complex (MHC) class II molecules into the endocytic pathway are poorly understood. For many proteins, access to endosomal compartments is regulated by cytosolically expressed sequences. We present evidence that a sequence in the lumenal domain of the MHC class II molecule regulates a very late event in class II biogenesis. Class II molecules containing single amino acid changes in the highly conserved 80–82 region of the β chain were introduced into invariant chain (Ii)-negative fibroblasts with wild-type α chain, and the derived transfectants were analyzed biochemically. Using an endosomal isolation technique, we have quantified the level of class II molecules expressed in endocytic compartments and found that in the absence of Ii, approximately 15% of total cellular class II molecules can be isolated from endosomal compartments. Mutation at position 80 enhances this localization, while changes at positions 81 and 82 ablate class II expression in endosomal compartments. In addition, we have evaluated whether the induced changes in intracellular distribution of class II molecules were due to alterations in early biosynthetic events, indicative of misfolding of the molecules, or to modulation of later trafficking events more likely to be a consequence of the modulation of a specific transport event. Despite the dramatic effects on endosomal localization induced by the mutations, early bio-synthetic events and maturation of class II were unaffected by the mutations. Collectively, our data argue that late trafficking events that control the ability of the class II molecule to access antigens is regulated by the 80–82 segment of the MHC class II β chains. 相似文献
4.
James M. Sheil Todd D. Schell Sara E. Shepherd Gerald F. Klimo John M. Kioschos Yvonne Paterson 《European journal of immunology》1994,24(9):2141-2149
In this study the immunogenic tryptic fragment from a horse cytochrome c (cyt c) digest recognized by cytotoxic T lymphocytes (CTL), induced by in vitro peptide stimulation from C57BL/6 (B6) and mutant B6.C-H-2bm1 (bm1) mice is identified. An identical sequence, p40—53, is recognized by CTL from both B6 and bm1 mice. In addition, both B6 and bm1 cloned CTL lines display unusual major histocompatibility complex (MHC) class I-restricted recognition of this peptide in that they respond to it in the context of H-2Kb, H-2Db, and H-2Kbm1 class I molecules, although the sequence lacks the usual structural Kb and Db peptide-binding motifs. Truncated analogues which resemble the lengths of naturally processed MHC class I-presented peptides, confer reactivity for B6 and bm1 CTL against EL4 (H-2b) targets as well as the L cell transfectants, L + Kb, L + Db, and L + Kbm1. The antigenic peptide with the greatest potency is p41—49, which appears to be generated by angiotensin converting enzyme cleavage of the full-length p40—53 tryptic peptide. The minimum antigenic peptide recognized by both B6 and bm1 CTL, and which targets lysis on each of the transfectants, is the hexamer p43—48 peptide from horse cyt c. Residues Pro44 and Thr47, which occupy polymorphic positions with respect to other species-variant cyt c molecules, influence recognition of these peptides differently for the B6 and bm1 CTL. The ability of H-2Kb, H-2Db, and mutant H-2Kbm1 class I molecules to present the same peptide to a single cloned CTL is discussed in the context of current knowledge of peptide anchor residues and side chain-specific binding pockets in the MHC class I peptide-binding site. 相似文献
5.
Lars
. Pedersen Anette Stryhn Thor L. Holtet Mikael Etzerodt Jens Gerwien Mogens H. Nissen Hans C. Thgersen Seren Buus 《European journal of immunology》1995,25(6):1609-1616
The function of major histocompatibility complex (MHC) class I molecules is to sample peptides derived from intracellular proteins and to present these peptides to CD8+ cytotoxic T lymphocytes. In this paper, biochemical assays addressing MHC class I binding of both peptide and β2-microglobulin (β2m) have been used to examine the assembly of the trimolecular MHC class I/β2m/peptide complex. Recombinant human β2m and mouse β2m2 have been generated to compare the binding of the two β2m to mouse class I. It is frequently assumed that human β2m binds to mouse class I heavy chain with a much higher affinity than mouse β2m itself. We find that human β2m only binds to mouse class I heavy chain with slightly (about 3-fold) higher affinity than mouse β2m. In addition, we compared the effect of the two β2m upon peptide binding to mouse class I. The ability of human β2m to support peptide binding correlated well with its ability to saturate mouse class I heavy chains. Surprisingly, mouse β2m only facilitated peptide binding when mouse β2m was used in excess (about 20-fold) of what was needed to saturate the class I heavy chains. The inefficiency of mouse β2m to support peptide binding could not be attributed to a reduced affinity of mouse β2m/MHC class I complexes for peptides or to a reduction in the fraction of mouse β2m/MHC class I molecules participating in peptide binding. We have previously shown that only a minor fraction of class I molecules are involved in peptide binding, whereas most of class I molecules are involved in β2m binding. We propose that mouse β2m interacts with the minor peptide binding (i.e. the “empty”) fraction with a lower affinity than human β2m does, whereas mouse and human β2m interact with the major peptide-occupied fraction with almost similar affinities. This would explain why mouse β2m is less efficient than human β2m in generating the peptide binding moiety, and identifies the empty MHC class I heavy chain as the molecule that binds human β2m preferentially. 相似文献
6.
Ennio Carbone Gyrgy Stuber Sofia Andre Lars Franksson Eva Klein Alberto Beretta Antonio G. Siccardi Klas Krre 《European journal of immunology》1993,23(8):1752-1756
Enhancement of major histocompatibility complex (MHC) class I expression leads to protection from recognition by natural killer (NK) cells in several systems. MHC class I gene products can be expressed in different forms at the cell surface - for example as “empty” β2-microglobulin (β2m)-associated heterodimers or free heavy chains. To study the role of different class I heavy chain forms in NK target interactions, we have used lymphoblastoid target cell lines preincubated with β2m. This was found to shift the equilibrium between β2m-associated and nonassociated - heavy chains in favor of the former. In parallel, there was a significant increase in NK sensitivity. The recognition of MHC class I-deficient cell lines was not affected by β2m, arguing against a general nonspecific effect of fern on NK sensitivity. Our data indicate that protection against NK recognition correlates with target cell expression of free heavy chains (i.e. devoid of β2m) rather than with expression of complexes. 相似文献
7.
Katinka M. Smits Peter J. K. Kuppen Alexander M. M. Eggermont Takuya Tamatani Masayuki Miyasaka Gert Jan Fleuren 《European journal of immunology》1994,24(1):171-175
The precise mechanism by which target cells are recognized and subsequently lysed by interleukin-2-activated natural killer (A-NK) cells is poorly understood. In this study the role of major histocompatibility complex (MHC) class I and adhesion molecules in the recognition and lysis of tumor cells was investigated in a syngeneic Wag rat model. Preincubation of tumor cells with F(ab′)2 fragments of anti-MHC class I monoclonal antibody (mAb) OX18 strongly enhanced the A-NK cell-mediated lysis. Also normal syngeneic cells such as T cells and A-NK cells became highly sensitive for lysis by A-NK cells after preincubation with mAb OX18. Two other mAb against MHC class I had no effect on lysis of target cells. These data indicate that masking of MHC class I on syngeneic tumor and normal cells by mAb OX18 is sufficient for A-NK cells to recognize target cells as non-self, resulting in lysis. In addition, we found that the presence of mAb against the β2 (CD18)-integrins blocked the lysis of all tumor cell lines by A-NK cells in 51Cr-release assays, also when target cells were preincubated with mAb OX18. Because of the absence of CD18 on most tumor cells we concluded that a CD18-associated integrin on A-NK cells is essential for lysis of target cells. These results show that in this syngeneic rat model CD18 on A-NK cells together with MHC class I on tumor cells determine A-NK cell-mediated lysis. Furthermore, we hypothesize that the anti-MHC class I OX18 recognizes an epitope on rat MHC class I which is, or is very close to, the restriction element determining A-NK cell-mediated lysis. 相似文献