摘要
Previous work from this laboratory has demonstrated that the addition of angiotensin(Aug)Ⅱresults in the rapid transcriptional activation of early growth response gene c-fos.Blockage of this increase completely inhibits the Aug Ⅱinduced increase in vascular smooth muscle cell(VSMC)growth.To explore the molecular mechanism responsible for the induction of c-fos in VSMC,a series of constructs containing portions of c-fos promoter linked to the reporter gene chloramphenicol acetyltransferase(CAT)were used in transient transfection assays.When a construct containing both the well described serum response element(SRE)and the cyclic AMP response element(CRE)was used,no endogenous CAT activity was observed in serum starved cells.The addition of either Ang Ⅱor serum resulted in a marked increase in CAT activity.Mutations in either the SRE or CRE alone which have been demonstrated to inactivate these elements in number of cell types had no effect on c-fos inducibility by either Angll or serum. In contrast,if both elements were mutated in the same construct,inducibility was reduced by 75 %￣80%.Using a construct in which the SRE has been deleted,a mutation in the CRE completely abolished induction of c-fos by either Aug Ⅱor serum. Mobility shift assays demonstrated that tow proteins bind specifically to the SRE and three proteins to CRE. These data demonstrate that the induction of c-fos in VSMC's can be mediated by two distinct enhancer elements each of which can act independently. Future research will be aimed at identifying the proteins that interact with these elemetns delineating the mechanisms by which Ang Ⅱstimulates their activity.
Previous work from this laboratory has demonstrated that the addition of angiotensin(Aug)Ⅱresults in the rapid transcriptional activation of early growth response gene c-fos.Blockage of this increase completely inhibits the Aug Ⅱinduced increase in vascular smooth muscle cell(VSMC)growth.To explore the molecular mechanism responsible for the induction of c-fos in VSMC,a series of constructs containing portions of c-fos promoter linked to the reporter gene chloramphenicol acetyltransferase(CAT)were used in transient transfection assays.When a construct containing both the well described serum response element(SRE)and the cyclic AMP response element(CRE)was used,no endogenous CAT activity was observed in serum starved cells.The addition of either Ang Ⅱor serum resulted in a marked increase in CAT activity.Mutations in either the SRE or CRE alone which have been demonstrated to inactivate these elements in number of cell types had no effect on c-fos inducibility by either Angll or serum. In contrast,if both elements were mutated in the same construct,inducibility was reduced by 75 %￣80%.Using a construct in which the SRE has been deleted,a mutation in the CRE completely abolished induction of c-fos by either Aug Ⅱor serum. Mobility shift assays demonstrated that tow proteins bind specifically to the SRE and three proteins to CRE. These data demonstrate that the induction of c-fos in VSMC's can be mediated by two distinct enhancer elements each of which can act independently. Future research will be aimed at identifying the proteins that interact with these elemetns delineating the mechanisms by which Ang Ⅱstimulates their activity.
