The Carboxyl-terminal Fragment of Pro-HB-EGF Reverses Bcl6-mediated Gene Repression

Masatsugu Hori,Yumi Kinugasa,Miki Hieda,Shigeki Higashiyama

doi:10.1074/jbc.m611036200

Masatsugu Hori, Yumi Kinugasa + Show 2 more

Open Access

https://doi.org/10.1074/jbc.m611036200

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Abstract

Heparin-binding epidermal growth factor-like growth factor (HB-EGF), a member of the EGF family, is synthesized as a type I transmembrane precursor (pro-HB-EGF). Ectodomain shedding of pro-HB-EGF yields an amino-terminal soluble ligand of EGF receptor (HB-EGF) and a carboxyl-terminal fragment (HB-EGF-CTF) consisting of the transmembrane and cytoplasmic domains. We previously showed that the HB-EGF-CTF translocates from the plasma membrane to the nucleus and plays a role as a signaling molecule. Immunoprecipitation showed that HB-EGF-CTF can associate with Bcl6, a transcriptional repressor in mammalian cells. A glutathione S-transferase pulldown assay revealed that HB-EGF-CTF interacted efficiently with zinc fingers 4-6 of Bcl6. A luciferase reporter assay showed that the nuclear translocation of HB-EGF-CTF following shedding reversed transcriptional repression of cyclin D2 by Bcl6. Additionally, the level of cyclin D2 protein increased and Bcl6 interaction with the cyclin D2 promoter decreased in parallel with the shedding of pro-HB-EGF at all endogenous levels. These findings suggest that HB-EGF-CTF is a potent regulator of gene expression via its interaction with the transcriptional repressor Bcl6.

Highlights

Heparin-binding EGF2-like growth factor (HB-EGF), a member of the EGF family, directly binds to and activates the EGFR (ErbB1/HER1) and can indirectly transactivate ErbB2 (HER2/ neu), ErbB3 (HER3), and ErbB4 (HER4) by forming a heterodimer with the EGFR [1]
We showed that HB-EGF-CTF is translocated from the plasma membrane to the nucleus and binds to the promyelocytic leukemia zinc finger (PLZF) protein, which promotes entry into the S-phase of the cell cycle [17,18,19]
Antibodies against the cytoplasmic region of pro-HB-EGF (#H1) immunoprecipitated APtagged pro-HB-EGF (Fig. 1A, lower) and an ϳ90-kDa band from the cell lysates. This corresponds to FLAG-B-cell lymphoma 6 (Bcl6), indicating that HB-EGF-CTF physically interacts with Bcl6 (Fig. 1A, upper)

Summary

Introduction

Heparin-binding EGF2-like growth factor (HB-EGF), a member of the EGF family, directly binds to and activates the EGFR (ErbB1/HER1) and can indirectly transactivate ErbB2 (HER2/ neu), ErbB3 (HER3), and ErbB4 (HER4) by forming a heterodimer with the EGFR [1]. A luciferase reporter assay showed that the nuclear translocation of HB-EGF-CTF following shedding reversed transcriptional repression of cyclin D2 by Bcl6. Our results indicate that HB-EGF-CTF interacts with Bcl6 and reverses the transcriptional repression of the cyclin D2 gene expression by Bcl6.

Results

Conclusion