Phosphorylated C/EBPβ Influences a Complex Network Involving YY1 and USF2 in Lung Epithelial Cells

Abstract

The promoter of the cystic fibrosis transmembrane conductance regulator gene CFTR is tightly controlled by regulators including CCAAT/enhancer binding proteins (C/EBPs). We previously reported that the transcription factors YY1 and USF2 affect CFTR expression. We can now demonstrate that C/EBPβ, a member of the CCAAT family, binds to the CFTR promoter and contributes to its transcriptional activity. Our data reveal that C/EBPβ cooperates with USF2 and acts antagonistically to YY1 in the control of CFTR expression. Interestingly, YY1, a strong repressor, fails to repress the CFTR activation induced by USF2 through DNA binding competition. Collectively, the data strongly suggest a model by which USF2 functionally interacts with YY1 blocking its inhibitory activity, in favour of C/EBPβ transactivation. Further investigation into the interactions between these three proteins revealed that phosphorylation of C/EBPβ influences the DNA occupancy of YY1 and favours the interaction between USF2 and YY1. This phosphorylation process has several implications in the CFTR transcriptional process, thus evoking an additional layer of complexity to the mechanisms influencing CFTR gene regulation.

The Basic Helix‐Loop‐Helix/Leucine Zipper Transcription Factor USF2 Integrates Serum‐Induced PAI‐1 Expression and Keratinocyte Growth

“…YY1, in fact, inhibits SMAD occupancy of the PAI-1 promoter likely via Interactions with the N-terminal MAD homology domain in SMAD4 and SMAD2/3 with suppression dependent on the number of SMAD-binding elements in the specific target gene [Kurisaki et al, 2003]. Importantly, USF2 appears to contribute to C/EBPβ target gene expression by interacting with YY1, either via the repressive domain or by blocking formation of YY1/co-regulator complexes to antagonize YY1 [Viart et al, 2013]. Interactions between USF2 and YY1 release YY1 inhibition stimulating C/EBPβ transactivation of target genes [Viart et al, 2013].…”

“…Importantly, USF2 appears to contribute to C/EBPβ target gene expression by interacting with YY1, either via the repressive domain or by blocking formation of YY1/co-regulator complexes to antagonize YY1 [Viart et al, 2013]. Interactions between USF2 and YY1 release YY1 inhibition stimulating C/EBPβ transactivation of target genes [Viart et al, 2013]. Other mechanisms have implications with regard to cell growth control.…”

Transcription factors and miRNAs that regulate fetal to adultCFTRexpression change are new targets for cystic fibrosis

“…Quantitative chromatin immunoprecipitation (Q-ChIP) was carried out as previously described [24]. Purified cross-linked chromatin was immunoprecipitated with 3 μg of each antibody (Santa Cruz, Heidelberg, Germany; Clinisciences, Montrouge, France).…”

“…For instance, CCAAT-enhancer-binding protein (C/EBP)δ positively regulates CFTR transcription by binding to an inverted CCAAT element in the promoter [16]. In addition, C/EBPβ binds to a DNase I-hypersensitive site that is present only in tissues expressing CFTR [23] and we recently demonstrated its binding to CFTR minimal promoter [24]. Chromatin structures that facilitate long-range interactions between various regulatory elements cluster specifically to the CFTR promoter exclusively in CFTR expressing cells [25,26].…”

Role of Non-coding RNAs in Cystic Fibrosis

“…Physiological CFTR expression is tightly controlled by transcriptional, post-transcriptional, translational and post-translational regulatory mechanisms, resulting in complex spatial and temporal expression patterns. Notwithstanding the importance of CFTR transcriptional regulation [1][2][3], CFTR expression can be modulated through other mechanisms. Indeed, epigenetic changes, such as DNA methylation or histone acetylation, also influence CFTR gene expression in different tissues [4][5][6][7].…”