역전사 중합효소 연쇄반응을 이용한 사람의 활성 T세포 항원 CTLA-4 유전자 클로닝 및 마우스 L세포에서의 발현 = Cloning of Human CTL-4 cDNA by Using Reverse Transcription-polymerase Chain Reaction and It's Expression in Murine L cell

다국어 초록 (Multilingual Abstract)

Occupancy of the T-cell receptor (TCR) by antigen in association with class Ⅱ MHC molecule is required for the initiation of T cell activation. However TCR stimulation is not a sufficient signal to account for all the observed events that occur during T cell activation. And in the absence of a second costimulatory signal provided by abtigen presenting cell antigen specific signal may lead to clonal inactivation or energy. CD28, a T cell surface molecule first defined by monoclonal antibody 9.3 on human cells, appears to function as an alternative, TCR/CD3-independent, activation pathway for T cell. This molecule provides costimulatory signal for T cell proliferation by increasing a number of lymphokines and cytokines production. A T cell activation antigen CTLA-4 is homologous to CD28 with 28% amino acid and 67% of nucleic acid homology in the protein coding region. CTLA-4 was ofiginally identified by the differential screening follosing subtractive hybridization of murine cytotoxic T cell cDNA library and mapped to the same lacation on chromosome 2 in human and chromosome 1 in mouse as CD28. These findings and activation associated expression pattern of CTLA-4 raise question about functional role of this molecule in T cell activation. In this study human CTLA-4 cDNA was amplified from phytohemagglutinin (PHA)-stimulated human peropheral blood lymphocytes mRNA by using reverse transcription-ploymerase chain reaction (RT-PCR) method. And amplified human CTLA-4 cDNA was subsequently cloned into pRC/CMV vector. In double stranded DNA sequencing the 331st base of protein coding region was changed from guanylic acid of human CTLA-4 from alanine to threonine. Murind L cell transfected with this clone expressed 36kD protein. It appears that this dirrerence of molecular weight between native (34kD) and recombinant (36kD) human CTLA-4 molecule may due to glycosylation differences. The cDNA and murine CTLA-4 molecule may due to glycosylation differences. The cDNA and murine L cell lines expressing human CTLA-4 developed by this study would contribute in future work unveiling the biological rile of CTLA-4 in T cell activation.