Identification of the linker histone H1 as a protein kinase C epsilon-binding protein in vascular smooth muscle

Zhao, Mingcai, Sutherland, Cindy, Wilson, David P., Deng, Jingti, MacDonald, Justin A. and Walsh, Michael P. (2004) Identification of the linker histone H1 as a protein kinase C epsilon-binding protein in vascular smooth muscle. Biochemistry And Cell Biology, 82 5: 538-546. doi:10.1139/O04-053

Author Zhao, Mingcai
Sutherland, Cindy
Wilson, David P.
Deng, Jingti
MacDonald, Justin A.
Walsh, Michael P.
Title Identification of the linker histone H1 as a protein kinase C epsilon-binding protein in vascular smooth muscle
Journal name Biochemistry And Cell Biology   Check publisher's open access policy
ISSN 0829-8211
Publication date 2004-10-01
Sub-type Article (original research)
DOI 10.1139/O04-053
Open Access Status
Volume 82
Issue 5
Start page 538
End page 546
Total pages 9
Place of publication Ottawa, ON, Canada
Publisher National Research Council Canada
Language eng
Subject 0913 Mechanical Engineering
Abstract A variety of anchoring proteins target specific protein kinase C (PKC) isoenzymes to particular subcellular locations or multimeric signaling complexes, thereby achieving a high degree of substrate specificity by localizing the kinase in proximity to specific substrates. PKCepsilon is widely expressed in smooth muscle tissues, but little is known about its targeting and substrate specificity. We have used a Far-Western (overlay) approach to identify PKCepsilon-binding proteins in vascular smooth muscle of the rat aorta. Proteins of similar to32 and 34 kDa in the Triton-insoluble fraction were found to bind PKCepsilon in a phospholipid/diacylglycerol-dependent manner. Although of similar molecular weight to RACK-1, a known PKCepsilon-binding protein, these proteins were separated from RACK-1 by SDS-PAGE and differential NaCl extraction and were not recognized by an antibody to RACK-1. The PKCepsilon-binding proteins were further purified from the Triton-insoluble fraction and identified by de novo sequencing of selected tryptic peptides by tandem mass spectrometry as variants of the linker histone H1. Their identity was confirmed by Western blotting with anti-histone H1 and the demonstration that purified histone H1 binds PKCepsilon in the presence of phospholipid and diacylglycerol but absence of Ca2+. The interaction of PKCepsilon with histone H1 was specific since no interaction was observed with histones H2A, H2S or H3S. Bound PKCepsilon phosphorylated histone H1 in a phospholipid/diacylglycerol-dependent but Ca2+-independent manner. Ca2+-dependent PKC was also shown to interact with histone H1 but not other histones. These results suggest that histone H1 is both an anchoring protein and a substrate for activated PKCepsilon and other PKC isoenzymes and likely serves to localize activated PKCs that translocate to the nucleus in the vicinity of specific nuclear substrates including histone H1 itself. Since PKC isoenzymes have been implicated in regulation of gene expression, stable interaction with histone H1 may be an important step in this process.
Keyword Biochemistry & Molecular Biology
Cell Biology
protein kinase C
histone H1
signaling complexes
smooth muscle
Ca2+-independent Isoforms
Intracellular Receptor
Nuclear Translocation
Anchoring Proteins
Filamentous Actin
Adapter Proteins
Phorbol Ester
Q-Index Code C1
Q-Index Status Provisional Code
Institutional Status Unknown

Document type: Journal Article
Sub-type: Article (original research)
Collections: Excellence in Research Australia (ERA) - Collection
School of Mechanical & Mining Engineering Publications
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Citation counts: TR Web of Science Citation Count  Cited 9 times in Thomson Reuters Web of Science Article | Citations
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Created: Sat, 26 Jan 2008, 02:22:59 EST