A critical role of the Thy28-MYH9 axis in B cell-specific expression of the Pax5 gene in chicken B cells

PLoS One. 2015 Jan 21;10(1):e0116579. doi: 10.1371/journal.pone.0116579. eCollection 2015.

Abstract

Accumulating evidence suggests that Pax5 plays essential roles in B cell lineage commitment. However, molecular mechanisms of B cell-specific expression of Pax5 are not fully understood. Here, we applied insertional chromatin immunoprecipitation (iChIP) combined with stable isotope labeling using amino acids in cell culture (SILAC) (iChIP-SILAC) to direct identification of proteins interacting with the promoter region of the endogenous single-copy chicken Pax5 gene. By comparing B cells with macrophage-like cells trans-differentiated by ectopic expression of C/EBPβ, iChIP-SILAC detected B cell-specific interaction of a nuclear protein, Thy28/Thyn1, with the Pax5 1A promoter. Trans-differentiation of B cells into macrophage-like cells caused down-regulation of Thy28 expression. Loss-of-function of Thy28 induced decrease in Pax5 expression and recruitment of myosin-9 (MYH9), one of Thy28-interacting proteins, to the Pax5 1A promoter. Loss-of-function of MYH9 also induced decrease in Pax5 expression. Thus, our analysis revealed that Thy28 is functionally required for B cell-specific expression of Pax5 via recruitment of MYH9 to the Pax5 locus in chicken B cells.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Avian Proteins / metabolism*
  • B-Lymphocytes / metabolism*
  • Cell Culture Techniques
  • Cell Line
  • Chickens / genetics*
  • Chickens / immunology
  • Chickens / metabolism
  • Chromatin Immunoprecipitation
  • Gene Expression Regulation
  • Macrophages / metabolism
  • Myosin Heavy Chains / metabolism*
  • Nuclear Proteins / metabolism*
  • PAX5 Transcription Factor / genetics*
  • PAX5 Transcription Factor / metabolism

Substances

  • Avian Proteins
  • Nuclear Proteins
  • PAX5 Transcription Factor
  • Thy28 protein, Gallus gallus
  • Myosin Heavy Chains

Grant support

This work was supported by the Tokyo Biochemical Research Foundation, Nakatani Foundation for Advancement of Measuring Technologies in Biomedical Engineering (T.F.), the Asahi Glass Foundation, the Uehara Memorial Foundation (H.F.), Takeda Science Foundation, the Kurata Memorial Hitachi Science and Technology Foundation (T.F. and H.F.), Adaptable & Seamless Technology Transfer Program through Target-driven R&D (A-STEP) by the Japan Science and Technology Agency (JST) (#AS242Z00789Q) (T.F.), Grant-in-Aid for Young Scientists (B) (#22710185 and #25830131) (T.F.), Grant-in-Aid for Scientific Research on Innovative Areas “Cell Fate” (#23118516) (T.F.), “The Genofield” (#23114707), “Transcription Cycle” (#25118512) (H.F.) from the Ministry of Education, Culture, Sports, Science and Technology of Japan, and Program for Combined Research Fields from Immunology Frontier Research Center, Osaka University (H.F.). The funders had no role in study design, data collection and analysis, decision to publish or preparation of the manuscript.