Expression of human cytomegalovirus immediate early protein IE1 in insect cells: splicing of RNA and recognition by CD4+ T-cell clones.
Recombinant baculoviruses containing the unspliced gene (Bac-IE1) and a truncated cDNA (Bac-EX4) of the immediate early protein 1 (IE1) of human cytomegalovirus (HCMV) were constructed. The recombinant proteins IE1 and EX4 were expressed in Sf 9 insect cells. Immunoblot analyses using a specific monoclonal antibody or human sera from HCMV seropositive subjects revealed that the IE1 protein had an apparent molecular mass of 71 kDa which was similar to that observed in both HCMV infected human fibroblasts and infected or transfected human astrocytoma cells. Furthermore, HCMV-specific CD4+ T cell clones proliferated in the presence of IE1 or of EX4 used as a control, and appropriate antigen presenting cells. Our data on the IE1 gene provide evidence that two introns can be properly spliced out in baculovirus infected insect cells. The expressed proteins should be useful in further studies on the immune response to the virus.
Davrinche
C
C
INSERM, Unité 100, CHU Purpan, Toulouse, France.
Pasquier
C
C
Cerutti
M
M
Serradell
L
L
Clément
D
D
Devauchelle
G
G
Michelson
S
S
Davignon
J L
JL
D000818:Animals; D000956:Antigens, Viral / Q000096:biosynthesis* / Q000302:isolation & purification; D001254:Astrocytoma; D001483:Base Sequence; D015704:CD4 Antigens / Q000378:metabolism*; D002460:Cell Line; D002999:Clone Cells; D003587:Cytomegalovirus / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D017874:Immediate-Early Proteins; D015151:Immunoblotting; D008969:Molecular Sequence Data; D009036:Moths; D009838:Oligodeoxyribonucleotides; D016133:Polymerase Chain Reaction; D012326:RNA Splicing; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000302:isolation & purification; D015183:Restriction Mapping; D016176:T-Lymphocyte Subsets / Q000276:immunology*; D014162:Transfection; D014407:Tumor Cells, Cultured; D014763:Viral Matrix Proteins / Q000096:biosynthesis
D000956:Antigens, Viral; D015704:CD4 Antigens; D017874:Immediate-Early Proteins; D009838:Oligodeoxyribonucleotides; D011994:Recombinant Proteins; D014763:Viral Matrix Proteins; C044582:immediate-early proteins, cytomegalovirus