The role of regulatory domains in maintaining autoinhibition in the multidomain kinase PKCα

Ruth Sommese; Michael Ritt; Carter J. Swanson; Sivaraj Sivaramakrishnan

doi:10.1074/jbc.M116.768457

The role of regulatory domains in maintaining autoinhibition in the multidomain kinase PKCα

Ruth Sommese, Michael Ritt, Carter J. Swanson, Sivaraj Sivaramakrishnan

Research output: Contribution to journal › Article › peer-review

10 Scopus citations

Abstract

Resolving the conformational dynamics of large multidomain proteins has proven to be a significant challenge. Here we use a variety of techniques to dissect the roles of individual protein kinase Cα (PKCα) regulatory domains in maintaining catalytic autoinhibition. We find that whereas the pseudosubstrate domain is necessary for autoinhibition it is not sufficient. Instead, each regulatory domain (C1a, C1b, and C2) appears to strengthen the pseudosubstrate-catalytic domain interaction in a nucleotide-dependent manner. The pseudosubstrate and C1a domains, however, are minimally essential for maintaining the inactivated state. Furthermore, disrupting known interactions between the C1a and other regulatory domains releases the autoinhibited interaction and increases basal activity. Modulating this interaction between the catalytic and regulatory domains reveals a direct correlation between autoinhibition and membrane translocation following PKC activation.

Original language	English (US)
Pages (from-to)	2873-2880
Number of pages	8
Journal	Journal of Biological Chemistry
Volume	292
Issue number	7
DOIs	https://doi.org/10.1074/jbc.M116.768457
State	Published - Feb 17 2017

Bibliographical note

Funding Information:
This work was supported in part by American Heart Association Scientist Development Grant 13SDG14270009 and National Institutes of Health Grants 1DP2 CA186752-01 and 1-R01-GM-105646-01-A1 (to S. S.). The authors declare that they have no conflicts of interest with the contents of this article. The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health. For cell translocation assays, work was done using the Nikon A1Rsi confocal fluorescence microscope at the University of Minnesota, University Imaging Centers. Time-resolved fluorescence decay measurements were taken at the Biophysical Technology Center at the University of Minnesota.

Publisher Copyright:
© 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

Publisher link

10.1074/jbc.M116.768457

Find It

Find It at U of M Libraries

Cite this

@article{d65eeaea93a64d6b8b99847cd23f6856,

title = "The role of regulatory domains in maintaining autoinhibition in the multidomain kinase PKCα",

abstract = "Resolving the conformational dynamics of large multidomain proteins has proven to be a significant challenge. Here we use a variety of techniques to dissect the roles of individual protein kinase Cα (PKCα) regulatory domains in maintaining catalytic autoinhibition. We find that whereas the pseudosubstrate domain is necessary for autoinhibition it is not sufficient. Instead, each regulatory domain (C1a, C1b, and C2) appears to strengthen the pseudosubstrate-catalytic domain interaction in a nucleotide-dependent manner. The pseudosubstrate and C1a domains, however, are minimally essential for maintaining the inactivated state. Furthermore, disrupting known interactions between the C1a and other regulatory domains releases the autoinhibited interaction and increases basal activity. Modulating this interaction between the catalytic and regulatory domains reveals a direct correlation between autoinhibition and membrane translocation following PKC activation.",

author = "Ruth Sommese and Michael Ritt and Swanson, {Carter J.} and Sivaraj Sivaramakrishnan",

note = "Funding Information: This work was supported in part by American Heart Association Scientist Development Grant 13SDG14270009 and National Institutes of Health Grants 1DP2 CA186752-01 and 1-R01-GM-105646-01-A1 (to S. S.). The authors declare that they have no conflicts of interest with the contents of this article. The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health. For cell translocation assays, work was done using the Nikon A1Rsi confocal fluorescence microscope at the University of Minnesota, University Imaging Centers. Time-resolved fluorescence decay measurements were taken at the Biophysical Technology Center at the University of Minnesota. Publisher Copyright: {\textcopyright} 2017 by The American Society for Biochemistry and Molecular Biology, Inc.",

year = "2017",

month = feb,

day = "17",

doi = "10.1074/jbc.M116.768457",

language = "English (US)",

volume = "292",

pages = "2873--2880",

journal = "Journal of Biological Chemistry",

issn = "0021-9258",

publisher = "American Society for Biochemistry and Molecular Biology Inc.",

number = "7",

}

TY - JOUR

T1 - The role of regulatory domains in maintaining autoinhibition in the multidomain kinase PKCα

AU - Sommese, Ruth

AU - Ritt, Michael

AU - Swanson, Carter J.

AU - Sivaramakrishnan, Sivaraj

N1 - Funding Information: This work was supported in part by American Heart Association Scientist Development Grant 13SDG14270009 and National Institutes of Health Grants 1DP2 CA186752-01 and 1-R01-GM-105646-01-A1 (to S. S.). The authors declare that they have no conflicts of interest with the contents of this article. The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health. For cell translocation assays, work was done using the Nikon A1Rsi confocal fluorescence microscope at the University of Minnesota, University Imaging Centers. Time-resolved fluorescence decay measurements were taken at the Biophysical Technology Center at the University of Minnesota. Publisher Copyright: © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

PY - 2017/2/17

Y1 - 2017/2/17

N2 - Resolving the conformational dynamics of large multidomain proteins has proven to be a significant challenge. Here we use a variety of techniques to dissect the roles of individual protein kinase Cα (PKCα) regulatory domains in maintaining catalytic autoinhibition. We find that whereas the pseudosubstrate domain is necessary for autoinhibition it is not sufficient. Instead, each regulatory domain (C1a, C1b, and C2) appears to strengthen the pseudosubstrate-catalytic domain interaction in a nucleotide-dependent manner. The pseudosubstrate and C1a domains, however, are minimally essential for maintaining the inactivated state. Furthermore, disrupting known interactions between the C1a and other regulatory domains releases the autoinhibited interaction and increases basal activity. Modulating this interaction between the catalytic and regulatory domains reveals a direct correlation between autoinhibition and membrane translocation following PKC activation.

AB - Resolving the conformational dynamics of large multidomain proteins has proven to be a significant challenge. Here we use a variety of techniques to dissect the roles of individual protein kinase Cα (PKCα) regulatory domains in maintaining catalytic autoinhibition. We find that whereas the pseudosubstrate domain is necessary for autoinhibition it is not sufficient. Instead, each regulatory domain (C1a, C1b, and C2) appears to strengthen the pseudosubstrate-catalytic domain interaction in a nucleotide-dependent manner. The pseudosubstrate and C1a domains, however, are minimally essential for maintaining the inactivated state. Furthermore, disrupting known interactions between the C1a and other regulatory domains releases the autoinhibited interaction and increases basal activity. Modulating this interaction between the catalytic and regulatory domains reveals a direct correlation between autoinhibition and membrane translocation following PKC activation.

UR - http://www.scopus.com/inward/record.url?scp=85013218931&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=85013218931&partnerID=8YFLogxK

U2 - 10.1074/jbc.M116.768457

DO - 10.1074/jbc.M116.768457

M3 - Article

C2 - 28049730

AN - SCOPUS:85013218931

SN - 0021-9258

VL - 292

SP - 2873

EP - 2880

JO - Journal of Biological Chemistry

JF - Journal of Biological Chemistry

IS - 7

ER -

The role of regulatory domains in maintaining autoinhibition in the multidomain kinase PKCα

Abstract

Bibliographical note

Publisher link

Find It

Other files and links

Fingerprint

Nikon A1RSI Confocal with SIM Super Resolution

University Imaging Centers

Cite this

The role of regulatory domains in maintaining autoinhibition in the multidomain kinase PKCα

Abstract

Bibliographical note

Publisher link

Find It

Other files and links

Fingerprint

University Assets

Nikon A1RSI Confocal with SIM Super Resolution

University Imaging Centers

Cite this