PARP-1 determines specificity in a retinoid signaling pathway via direct modulation of mediator.
We show that PARP-1 is indispensable to retinoic acid receptor (RAR)-mediated transcription from the RARbeta2 promoter in a highly purified, reconstituted transcription system and that RA-inducible expression of all RARbeta isoforms is abrogated in PARP-1(-/-) cells in vivo. Importantly, PARP-1 activity was independent of its catalytic domain. PARP-1 directly interacts with RAR and Mediator. Chromatin immunoprecipitation experiments confirmed the presence of PARP-1 and Mediator on RAR-responsive promoters in vivo. Importantly, Mediator was inactive (Cdk8+) under basal conditions but was activated (Cdk8-) upon induction. However, in PARP-1(-/-) cells, Mediator was retained in its inactive state (Cdk8+) upon induction consistent with the absence of gene expression. PARP-1 became dispensable for ligand-dependent transcription in a chromatin reconstituted transcription assay when Mediator was devoid of the Cdk8 module (CRSP). PARP-1 appears to function as a specificity factor regulating the RA-induced switch of Mediator from the inactive (Cdk8+) to the active (Cdk8-) state in RAR-dependent transcription.
Pavri
Rushad
R
Department of Biochemistry, Howard Hughes Medical Institute, University of Medicine and Dentistry of New Jersey, 683 Hoes Lane, Piscataway, New Jersey 08854, USA.
Lewis
Brian
B
Kim
Tae-Kyung
TK
Dilworth
F Jeffrey
FJ
Erdjument-Bromage
Hediye
H
Tempst
Paul
P
de Murcia
Gilbert
G
Evans
Ronald
R
Chambon
Pierre
P
Reinberg
Danny
D
D000818:Animals; D002460:Cell Line; D002467:Cell Nucleus / Q000235:genetics / Q000502:physiology; D019281:Dimerization; D017353:Gene Deletion; D006367:HeLa Cells; D006801:Humans; D011065:Poly(ADP-ribose) Polymerases / Q000235:genetics / Q000378:metabolism*; D018168:Receptors, Retinoic Acid / Q000235:genetics / Q000378:metabolism*; D011994:Recombinant Proteins / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D015398:Signal Transduction / Q000502:physiology*; D014158:Transcription, Genetic / Q000235:genetics; D014162:Transfection
D018168:Receptors, Retinoic Acid; D011994:Recombinant Proteins; C082925:retinoic acid receptor beta; D011065:Poly(ADP-ribose) Polymerases