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Molecular cloning of rat NK target structure--the possibility of CD44 involvement in NK cell-mediated lysis
1Department of Pathology, Sapporo Medical University School of Medicine, Hokkaido, Japan.
Abstract:
The nature of target molecules of natural killer (NK) cell-mediated lysis remains to be elucidated. As we previously reported, mAb 109 recognizes one of the tumor-associated antigens, designated as 109 antigen (Ag), expressed on the cell surface of rat fibrosarcomas W31 and W14, which are transformants of WFB (rat fetal fibroblast cell line) with H-ras oncogene. 109Ag was thought to be a target structure of NK cells since mAb 109 inhibited NK cell-mediated lysis against W31 and W14. Here, we demonstrate by molecular cloning that 109Ag is identical to rat CD44. Immunoprecipitation and immunoblotting studies also showed that mAb 109 and anti-rat CD44 mAb OX-50 recognize the same protein of W31 cell lysates with an 86 kDa molecular size. CD44 was suggested to be a target structure of NK cell-mediated lysis; however, rat CD44 cDNA transfection alone into CD44 null cell lines did not result in up-regulation of target cell susceptibility to NK cell-mediated lysis. Our results therefore indicated that CD44 may play a crucial role as one of the target structures in our rat fibrosarcoma system though the cell surface expression of CD44 alone does not affect NK susceptibility of the target cells.
Insights
Natural killer (NK) cell lysis targets were identified. Rat CD44 was found to be identical to 109 antigen, a tumor-associated antigen, but CD44 expression alone did not increase NK cell susceptibility.
Area of Science:
- Immunology
- Molecular Biology
- Oncology
Background:
- Natural killer (NK) cell-mediated lysis targets are not fully understood.
- Previous work identified 109 antigen (Ag) on rat fibrosarcomas as a potential NK cell target.
- Monoclonal antibody (mAb) 109 inhibited NK cell lysis of tumor cells expressing 109Ag.
Purpose of the Study:
- To elucidate the molecular identity of the 109 antigen.
- To determine if CD44 is a target structure for NK cell-mediated lysis in this fibrosarcoma model.
- To investigate the role of CD44 expression in NK cell susceptibility.
Main Methods:
- Molecular cloning to identify 109Ag.
- Immunoprecipitation and immunoblotting to characterize the protein recognized by mAb 109.
- Transfection of CD44 cDNA into CD44-null cell lines.
Main Results:
- Molecular cloning confirmed that 109Ag is identical to rat CD44.
- mAb 109 and anti-rat CD44 mAb OX-50 recognize the same 86 kDa protein.
- Transfection of rat CD44 cDNA alone into CD44-null cells did not increase susceptibility to NK cell lysis.
Conclusions:
- Rat CD44 is the 109 antigen and plays a role as a target structure in this rat fibrosarcoma system.
- Cell surface expression of CD44 alone is insufficient to confer susceptibility to NK cell-mediated lysis.
- Additional factors likely contribute to NK cell targeting in this model.
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