High Mobility Group Protein-B1 Interacts with Sterol Regulatory Element-binding Proteins to Enhance Their DNA Binding

Yuho Najima,Takashi Kadowaki,Nobuhiro Yamada,Hiroaki Okazaki,Sachiko Okazaki,Kenji Harada,Jun-Ichi Osuga,Hitoshi Shimano,Yoshimi Nakagawa,Motohiro Sekiya,Ryozo Nagai,Naoya Yahagi,Ken Ohashi,Yoshiaki Tamura,Takashi Matsuzaka,Yoshinori Takeuchi,Shun Ishibashi,Takanari Gotoda,Michiyo Amemiya-Kudo,Yoko Ikoma

doi:10.1074/jbc.m414549200

Yuho Najima, Takashi Kadowaki + Show 18 more

Open Access

https://doi.org/10.1074/jbc.m414549200

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Abstract

Sterol regulatory element-binding proteins (SREBPs) are transcription factors that are predominately involved in the regulation of lipogenic and cholesterogenic enzyme gene expression. To identify unknown proteins that interact with SREBP, we screened nuclear extract proteins with 35S-labeled SREBP-1 bait in Far Western blotting analysis. Using this approach, high mobility group protein-B1 (HMGB1), a chromosomal protein, was identified as a novel SREBP interacting protein. In vitro glutathione S-transferase pull-down and in vivo coimmunoprecipitation studies confirmed an interaction between HMGB1 and both SREBP-1 and -2. The protein-protein interaction was mediated through the helix-loop-helix domain of SREBP-1, residues 309-344, and the A box of HMGB1. Furthermore, an electrophoretic mobility shift assay demonstrated that HMGB1 enhances SREBPs binding to their cognate DNA sequences. Moreover, luciferase reporter analyses, including RNA interference technique showed that HMGB1 potentiates the transcriptional activities of SREBP in cultured cells. These findings raise the intriguing possibility that HMGB1 is potentially involved in the regulation of lipogenic and cholesterogenic gene transcription.

Highlights

Sterol regulatory element-binding proteins (SREBPs)1 are members of the basic helix-loop-helix leucine zipper (B-HLHZIP) family of transcription factors that bind to specific sterolresponsive elements (SREs) in the promoters of target genes and thereby regulate fatty acid and cholesterol synthesis
To further examine whether these proteins are coimmunoprecipitated with SREBP-1 using anti-SREBP-1 antibody, immunoprecipitated nuclear proteins were analyzed by Far Western technique in the same way
Homodimerization of SREBP-1 was detected at 66 kDa, which demonstrates that this Far Western blotting analysis is sensitive and specific with Western blotting

Summary

Introduction

Sterol regulatory element-binding proteins (SREBPs) are members of the basic helix-loop-helix leucine zipper (B-HLHZIP) family of transcription factors that bind to specific sterolresponsive elements (SREs) in the promoters of target genes and thereby regulate fatty acid and cholesterol synthesis (reviewed in Refs. 1–3). Sterol regulatory element-binding proteins (SREBPs) are members of the basic helix-loop-helix leucine zipper (B-HLHZIP) family of transcription factors that bind to specific sterolresponsive elements (SREs) in the promoters of target genes and thereby regulate fatty acid and cholesterol synthesis Sues, the predominant isoform is SREBP-1c, which controls gene expression of lipogenic enzymes, including fatty acid synthase and ATP citrate lyase, whereas SREBP-2 plays a crucial role in the regulation of key enzymes in the cholesterol synthetic pathway such as 3-hydroxy-3-methylglutaryl (HMG)CoA synthase and farnesyl diphosphate synthase (4 –9). HMGB1 is a chromatin-binding protein that belongs to high mobility group (HMG)-box family and appears to act as an architectural facilitator in the assembly of nucleoprotein complexes in a variety of DNA-related processes including transcription, replication, V(D)J recombination, and repair We describe the further function of HMGB1 in interacting with SREBPs

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