LHR gene transcription is subject to repression/derepression through various modes and multiple effectors. Epigenetic silencing and activation of the LHR is achieved through coordinated regulation at both histone and DNA levels. The LHR gene is subject to repression by deacetylation and methylation at its promoter region, where a HDAC/mSin3A co-repressor complex is anchored at Sp1 sites. The present studies revealed that PKC/ERK signaling is important for the activation of LHR promoter activity, and the increase of endogenous transcripts induced by phorbol-12-myristate-13-acetate (PMA) in HeLa cells. While these effects were attributable to PKC activity, the ERK pathway was the downstream effector in LHR activation. PMA caused a significant enhancement of Sp1 phosphorylation at serine residue (s), which was blocked by PKC or ERK inhibition. The interaction of activated phospho-ERK with Sp1 and ERK's association with the LHR promoter points to Sp1 as a direct target of ERK. Following Sp1 phosphorylation the HDAC1/mSin3A repressor complex dissociated from Sp1 sites, histone 3 was acetylated, and TFIIB and Pol II were recruited. In addition, overexpression of a constitutively active PKC (PKC CA) strongly activated LHR transcription in MCF-7 cells (devoid of PKC), induced Sp1 phosphorylation at serine residue (s), and caused derecruitment of HDAC1/mSin3A complex from the promoter. These effects were negated by cotransfection of a dominant negative PKC. In conclusion, these studies have revealed a novel regulatory signaling mechanism of transcriptional control in which the LHR is derepressed through PKC/ERK-mediated Sp1 phosphorylation, causing the release of HDAC1/mSin3A complex from the promoter.