RNA-Seq and ChIP-Seq reveal SQSTM1/p62 as a key mediator of JunB suppression of NF-κB-dependent inflammation

J Invest Dermatol. 2015 Apr;135(4):1016-1024. doi: 10.1038/jid.2014.519. Epub 2014 Dec 16.

Abstract

Mice with epidermal deletion of JunB transcription factor displayed a psoriasis-like inflammation. The relevance of these findings to humans and the mechanisms mediating JunB function are not fully understood. Here we demonstrate that impaired JunB function via gene silencing or overexpression of a dominant negative mutant increased human keratinocyte cell proliferation but decreased cell barrier function. RNA-seq revealed over 500 genes affected by JunB loss of function, which included the upregulation of an array of proinflammatory molecules relevant to psoriasis. Among these were tumor necrosis factor α (TNFα), CCL2, CXCL10, IL6R, and SQSTM1, an adaptor protein involved in nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) activation. Chromatin immunoprecipitation (ChIP)-Seq and gene reporter analyses showed that JunB directly suppressed SQSTM1 by binding to a consensus AP-1 cis element located around 2 kb upstream of SQSTM1-transcription start site. Similar to JunB loss of function, SQSTM1-overexpression induced TNFα, CCL2, and CXCL10. Conversely, NF-κB inhibition genetically with a mutant IκBα or pharmacologically with pyrrolidine dithiocarbamate (PDTC) prevented cytokine, but not IL6R, induction by JunB deficiency. Taken together, our findings indicate that JunB controls epidermal growth, barrier formation, and proinflammatory responses through direct and indirect mechanisms, pinpointing SQSTM1 as a key mediator of JunB suppression of NF-κB-dependent inflammation.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Adaptor Proteins, Signal Transducing / genetics*
  • Adaptor Proteins, Signal Transducing / metabolism
  • Animals
  • Cell Adhesion
  • Cell Proliferation
  • Chemokine CCL2 / metabolism
  • Chemokine CXCL10 / metabolism
  • Chromatin Immunoprecipitation
  • Cytokines / metabolism
  • Epidermis / metabolism
  • Gene Deletion
  • Genes, Reporter
  • HEK293 Cells
  • Heat-Shock Proteins / genetics*
  • Heat-Shock Proteins / metabolism
  • Humans
  • Inflammation / metabolism*
  • Keratinocytes / cytology
  • Mice
  • Mutation
  • NF-kappa B / metabolism*
  • Oligonucleotide Array Sequence Analysis
  • Pyrrolidines / chemistry
  • Regeneration
  • Sequence Analysis, RNA
  • Sequestosome-1 Protein
  • Skin Physiological Phenomena
  • Thiocarbamates / chemistry
  • Transcription Factors / genetics
  • Transcription Factors / metabolism*

Substances

  • Adaptor Proteins, Signal Transducing
  • Ccl2 protein, mouse
  • Chemokine CCL2
  • Chemokine CXCL10
  • Cxcl10 protein, mouse
  • Cytokines
  • Heat-Shock Proteins
  • JunB protein, mouse
  • NF-kappa B
  • Pyrrolidines
  • Sequestosome-1 Protein
  • Sqstm1 protein, mouse
  • Thiocarbamates
  • Transcription Factors
  • pyrrolidine dithiocarbamic acid