Lack of an absolute requirement for the native aryl hydrocarbon receptor (AhR) and AhR nuclear translocator transactivation domains in protein kinaseC-mediated modulation of the AhR pathway
Wp. Long et Gh. Perdew, Lack of an absolute requirement for the native aryl hydrocarbon receptor (AhR) and AhR nuclear translocator transactivation domains in protein kinaseC-mediated modulation of the AhR pathway, ARCH BIOCH, 371(2), 1999, pp. 246-259
Protein kinase C (PKC)-mediated modulation of the aryl hydrocarbon receptor
(AhR) pathway was examined in CROK1-derived L10.I cells stably transfected
with the pGUDLUC6.1 reporter; pGUDLUC6.1 is solely controlled by four diox
in-responsive enhancer elements, Co treatment of L10.I cells with 10 nM 2,3
,7,8-tetrachlorodibenzo-p-dioxin (TCDD) and 81 nM phorbol 12-myristate 13-a
cetate (PMA), an activator of sn-l,2-diacylglyerol binding PKCs, enhanced t
ransactivation of the reporter construct several-fold relative to cells tre
ated with a saturating 10 nM TCDD dose alone; this effect was dubbed the "P
MA effect." A domain swapping and deletional analysis of the native AhR and
AhR nuclear translocator (ARNT) protein transactivation domains (TADs) was
performed to determine if these domains are absolutely required for the Ah
R.ARNT dimer-mediated PMA effect in the L10.I model system; controls demons
trate the suitability of the L10.I model for these analyses and that endoge
nous AhR and ARNT levels are extremely low in this model. Transient coexpre
ssion of the AhR and ARNT-474-FLAG, an ARNT protein lacking the native ARNT
TAD, in L10.I cells reveals the native ARNT TAD is not absolutely required
for the AhR.ARNT-474-FLAG dimer to mediate the PMA effect. Transient coexp
ression of AhR Delta CVP, a chimeric AhR protein in which the native AhR TA
D has been replaced with the VP16 (herpes simplex virus protein IB) TAD (wh
ich control experiments demonstrate is unaffected by PMA), and ARNT in L10.
I cells indicates that the native AhR TAD is not absolutely required for th
is AhR Delta CVP.ARNT dimer to mediate the PMA effect, These observations s
trongly suggest that PKC-mediated modulation of the AhR pathway is not abso
lutely dependent on coactivators recruited to the AhR.ARNT dimer by the nat
ive TADs of the AhR and its heterodimerization partner ARNT. (C) 1999 Acade
mic Press.