Fig. 2. Transactivation of the deletion mutants of the TGF-ß RII promoter by
v-Src. A, schematic representation of TGF-ß RII
promoter deletion is shown. Serial deletion mutants of the 5'-flanking
region of the TGF-ß RII gene were
ligated to luciferase gene (pGL2-basic vector) and
cotransfected with v-Src expression vector (pMvsrc) into HepG2 cell.
B, pTßRIIP-219/+2-CAT and pTßRIIP+2/+36-CAT, which
contain v-src-responsive elements, were cotransfected with v-Src
expression vector into the 32D-123 cell line, and cells were harvested
after 48 h. The results show a representative CAT assay.
C, a series of deletion mutants from the regions -219
to +36 of the RII gene were subcloned into the
promoterless pGL3-basic vector and transiently cotransfected with v-Src
expression vector into HepG2 cell. Numbers indicate the position
relative to transcription start site. D, to characterize
other v-Src-responsive elements except for the inverted CCAAT box, the
inverted CCAAT region was deleted from pTßRIIP-219/+36 by
PvuII restriction enzyme, generating
pTßRIIP-130/-8-luc. This construct was cotransfected with v-Src
expression vector into HepG2 cell. Results from these experiments are
averages of at least triplicate experiments of three independent
transfections. Transfection efficiencies were normalized by ß-gal
activity as described in "Materials and Methods."