Liver-enriched Inhibitory Protein (LIP) Actively Inhibits Preadipocyte Differentiation through Histone Deacetylase 1 (HDAC1)

Abstract

The CCAAT/enhancer-binding protein ␤ (C/EBP␤) is expressed as three isoforms (LAP*, liver-enriched activating protein (LAP), and liver-enriched inhibitory protein (LIP)) that differentially regulate gene expression. The interplay between LAP*, LAP, and LIP in regulating cellular processes is largely unknown, and LIP has been largely regarded to repress transcription through a passive heterodimerization-dependent mechanism. Recently, we have shown that p300/GCN5 and mSin3A/HDAC1 differentially regulate the ability of C/EBP␤ to stimulate preadipocyte differentiation through activation of C/ebp␣ transcription. Here, we have mapped requirements for binding of mSin3A/HDAC1 to LAP/LAP* and LIP to a 4-amino acid motif in the central region of LAP/LAP* (residues 153-156) and the N terminus of LIP. Reducing mSin3A/HDAC1 binding to LAP/LAP* and LIP through deletion of this motif reduced the recruitment of HDAC1 to the C/ebp␣ promoter and increased preadipocyte differentiation stimulated by insulin and 1-methyl-3-isobutylxanthine. Additional studies showed that the interaction of HDAC1 with LIP provides for active repression of C/ebp␣ transcription and is largely responsible for the ability of LIP and HDAC1 to repress preadipocyte differentiation. Thus, although mSin3A/HDAC1 interacted readily with LAP/LAP* in addition to LIP and that expression of LAP/LAP* was sufficient to recruit HDAC1 to the C/ebp␣ promoter, mutations in C/ebp␤ that abrogated HDAC1 association to LAP/LAP* in the absence of LIP provided no additional stimulation of differentiation or transcription beyond the deletion of LIP alone. The implication of these results for the interaction between p300/GCN5 and mSin3A/HDAC1 in regulating C/EBP␣ transcription and preadipocyte differentiation are discussed.The CCAAT/enhancer-binding protein ␤ (C/EBP␤) 2 is a basic leucine zipper (bZip) transcription factor that plays key roles in cellular differentiation, proliferation, stress responses, and inflammation. The C/ebp␤ gene encodes three isoforms, LAP* (liver-enriched activating protein*), LAP, and LIP (liverenriched inhibitory protein) translated from a single mRNA from three consecutive in-frame methionines (Fig. 1A) (1, 2). Relative expression of LAP*/LAP/LIP varies according to cell type, and the three isoforms dimerize interchangeably and heterodimerize broadly with other bZip factors. The shorter LIP isoform is distinguished by the absence of the transcriptional activation functions that occur in the N terminus of LAP*/LAP. Thus, LIP has been primarily hypothesized to be a passive repressor of transcription that reduces transcriptional activation through competition for DNA-binding sites and the formation of heterodimers in which only one partner contributes to activation (1). Although the relative level of LAP/LAP* in a cell is regulatable, generally the level of LIP expression is approximately equivalent to the level of LAP, with LAP* levels usually being significantly lower (2). C/EBP␤ has been shown to play an important role at the onset of the differenti...

C/EBPβ and C/EBPδ transcription factors: Basic biology and roles in the CNS

“…C/EBPβ interacts with mSin3a/HDAC through a small AAGF motif between CR6 and CR7. This motif is located in the Nterminal extreme of LIP and mSin3a/HDAC recruitment is in fact an important mechanism of transcription inhibition by LIP (Abdou et al 2011). The association between C/EBPβ and mSin3a/HDAC1 can be decreased by C/EBPβ acetylation at lysines K98, K101 and K102 (Wiper-Bergeron et al 2007).…”

“…C/EBPδ has also been able to recruit and transactivate through CBP/p300 A c c e p t e d M a n u s c r i p t 25 (Chamberlain et al 2012;Wang et al 2006), residues Leu81 and Phe82 of C/EBPδ being critical for this interaction (Kovacs et al 2003) C/EBPβ also interacts with the complex formed by the co-repressor mSin3a and the histone deacetylase HDAC. This interaction, particularly with HDAC1, leads to repressed transcription of many genes, including C/EBPα (Abdou et al 2011;WiperBergeron et al 2003), p53, sirtuin 1 and glycogen synthase kinase 3β GSK3β Jin et al 2011;Jin et al 2009), PPARβ (Di-Poi et al 2005 or microglial CD200R1 (Dentesano et al 2012). Interaction of C/EBPδ with mSin3a/HDAC also inhibits transcription (Turgeon et al 2008).…”

Inactivation of Histone Deacetylase 1 (HDAC1) But Not HDAC2 Is Required for the Glucocorticoid-Dependent CCAAT/Enhancer-Binding Protein α (C/EBPα) Expression and Preadipocyte Differentiation

“…Glucocorticoids have been shown to affect the adipocyte differentiation process by up-regulating the expression of C/EBP␦, down-regulation of preadipocyte factor 1, up-regulation of transcription factors such as Forkhead box protein O1 and Kruppel-like factor 15, and through the up-regulation of C/EBP␣ (6 -11). We have reported that Dex treatment activated the transcriptional activity of C/EBP␤ at the C/ebp␣ promoter and increased adipogenesis (12)(13)(14)(15). Furthermore, it has been shown by us and others that the transcriptional potential of C/EBP␤ at the C/ebp␣ promoter is regulated by its acetylation status on critical residues (12,16) and its ability to release the inhibitory mammalian switch-independent 3A (mSin3A)/histone deacetylase (HDAC)1 complex through important regulatory domains (12,14,15).…”

“…Mutational analyses performed by other groups have demonstrated that substitutions of these amino acids compromise the ability of HDAC1 to deacetylate its substrates (22,30,31). HDAC1 has been implicated in a number of transcriptional processes and has been shown to interact directly and indirectly with a number of transcription factors such as p53, retinoblastoma protein, and C/EBP␤ and C/EBP␦ (13,14,(32)(33)(34). The role of HDAC1 in adipogenesis is somewhat controversial.…”

“…We have reported that Dex treatment activated the transcriptional activity of C/EBP␤ at the C/ebp␣ promoter and increased adipogenesis (12)(13)(14)(15). Furthermore, it has been shown by us and others that the transcriptional potential of C/EBP␤ at the C/ebp␣ promoter is regulated by its acetylation status on critical residues (12,16) and its ability to release the inhibitory mammalian switch-independent 3A (mSin3A)/histone deacetylase (HDAC)1 complex through important regulatory domains (12,14,15). A second member of the C/EBP family of transcription factor, C/EBP␦, has also been shown to be regulated by an mSin3A/HDAC1 corepressor complex that is sensitive to glucocorticoids (17).…”