Expression and characterization of mouse cFos protein using the baculovirus expression system: ability to form functional AP-1 complex with coexpressed cJun protein.
The products of the proto-oncogenes c-fos and c-jun play important roles in cell growth control, differentiation, and malignant transformation. Purified oncogenic proteins are essential tools in cell growth control studies. Here we describe the production of mouse cFos and cJun oncoproteins in Sf9 insect cells using the baculovirus expression system. The mouse c-fos cDNA was subcloned into two different baculovirus expression vectors, namely pVL1392 and pVLMH6, to generate, respectively, nonfusion and His-fusion cFos oncoproteins in insect cells. The products were characterized by immunofluorescence, immunoblotting, immunoprecipitation, and ability to bind to the in vitro translated JunB protein to form the AP-1 complex. A His-cJun fusion protein was also produced in insect cells using the pVLMH6 baculovirus vector. Coexpression of cFos and cJun in insect cells yielded functional AP-1 complexes as judged from gel retardation assays. The results point to the possibility of using insect cells for structural and functional studies of different Fos/Jun AP-1 complexes.
Corvello
C M
CM
Instituto de Química, Universidade de São Paulo, Brasil.
Metz
R
R
Bravo
R
R
Armelin
M C
MC
D000818:Animals; D016367:Baculoviridae; D001483:Base Sequence; D015139:Blotting, Southern; D003001:Cloning, Molecular; D004247:DNA / Q000378:metabolism; D018076:DNA, Complementary / Q000302:isolation & purification; D006801:Humans; D051379:Mice; D008969:Molecular Sequence Data; D011485:Protein Binding; D016760:Proto-Oncogene Proteins c-fos / Q000235:genetics*; D016755:Proto-Oncogene Proteins c-jun / Q000235:genetics*; D011993:Recombinant Fusion Proteins; D018808:Transcription Factor AP-1 / Q000378:metabolism*
D018076:DNA, Complementary; D016760:Proto-Oncogene Proteins c-fos; D016755:Proto-Oncogene Proteins c-jun; D011993:Recombinant Fusion Proteins; D018808:Transcription Factor AP-1; D004247:DNA