Conserved domains subserve novel mechanisms and functions in DKF-1, a Caenorhabditis elegans protein kinase D

Hui Feng, Min Ren, Charles S. Rubin

Research output: Contribution to journalArticle

13 Citations (Scopus)

Abstract

Protein kinase D (PKD) isoforms are effectors in signaling pathways controlled by diacylglycerol. PKDs contain conserved diacylglycerol binding (C1a, C1b), pleckstrin homology (PH), and Ser/Thr kinase domains. However, the properties of conserved domains may vary within the context of distinct PKD polypeptides. Such functional/structural malleability (plasticity) was explored by studying Caenorhabditis elegans D kinase family-1 (DKF-1), a PKD that governs locomotion in vivo. Phorbol ester binding with C1b alone activates classical PKDs by relieving C1-mediated inhibition. In contrast, C1a avidly ligated phorbol 12-myristate 13-acetate (PMA) and anchored DKF-1 at the plasma membrane. C1b bound PMA (moderate affinity) and cooperated with C1a in targeting DKF-1 to membranes. Mutations at a "Pro11" position in C1 domains were inactivating; kinase activity was minimal at PMA concentrations that stimulated wild type DKF-1 ∼10-fold. DKF-1 mutants exhibited unchanged, maximum kinase activity after cells were incubated with high PMA concentrations. Titration in situ revealed that translocation and activation of wild type and mutant DKF-1 were tightly and quantitatively linked at all PMA concentrations. Thus, C1 domains positively regulated phosphotransferase activity by docking DKF-1 with pools of activating lipid. A PH domain inhibits kinase activity in classical PKDs. The DKF-1 PH module neither inhibited catalytic activity nor bound phosphoinositides. Consequently, the PH module is an obligatory, positive regulator of DKF-1 activity that is compromised by mutation of Lys298 or Trp396. Phosphorylation of Thr588 switched on DKF-1 kinase activity. Persistent phosphorylation of Thr588 (activation loop) promoted ubiquitinylation and proteasome-mediated degradation of DKF-1. Each DKF-1 domain displayed novel properties indicative of functional malleability (plasticity).

Original languageEnglish (US)
Pages (from-to)17815-17826
Number of pages12
JournalJournal of Biological Chemistry
Volume281
Issue number26
DOIs
StatePublished - Jun 30 2006

Fingerprint

Caenorhabditis elegans Proteins
Phosphotransferases
Acetates
protein kinase D
Phosphorylation
Diglycerides
Plasticity
Chemical activation

ASJC Scopus subject areas

  • Biochemistry

Cite this

Conserved domains subserve novel mechanisms and functions in DKF-1, a Caenorhabditis elegans protein kinase D. / Feng, Hui; Ren, Min; Rubin, Charles S.

In: Journal of Biological Chemistry, Vol. 281, No. 26, 30.06.2006, p. 17815-17826.

Research output: Contribution to journalArticle

@article{b72af751ea9f4e34a4c339059284c02c,
title = "Conserved domains subserve novel mechanisms and functions in DKF-1, a Caenorhabditis elegans protein kinase D",
abstract = "Protein kinase D (PKD) isoforms are effectors in signaling pathways controlled by diacylglycerol. PKDs contain conserved diacylglycerol binding (C1a, C1b), pleckstrin homology (PH), and Ser/Thr kinase domains. However, the properties of conserved domains may vary within the context of distinct PKD polypeptides. Such functional/structural malleability (plasticity) was explored by studying Caenorhabditis elegans D kinase family-1 (DKF-1), a PKD that governs locomotion in vivo. Phorbol ester binding with C1b alone activates classical PKDs by relieving C1-mediated inhibition. In contrast, C1a avidly ligated phorbol 12-myristate 13-acetate (PMA) and anchored DKF-1 at the plasma membrane. C1b bound PMA (moderate affinity) and cooperated with C1a in targeting DKF-1 to membranes. Mutations at a {"}Pro11{"} position in C1 domains were inactivating; kinase activity was minimal at PMA concentrations that stimulated wild type DKF-1 ∼10-fold. DKF-1 mutants exhibited unchanged, maximum kinase activity after cells were incubated with high PMA concentrations. Titration in situ revealed that translocation and activation of wild type and mutant DKF-1 were tightly and quantitatively linked at all PMA concentrations. Thus, C1 domains positively regulated phosphotransferase activity by docking DKF-1 with pools of activating lipid. A PH domain inhibits kinase activity in classical PKDs. The DKF-1 PH module neither inhibited catalytic activity nor bound phosphoinositides. Consequently, the PH module is an obligatory, positive regulator of DKF-1 activity that is compromised by mutation of Lys298 or Trp396. Phosphorylation of Thr588 switched on DKF-1 kinase activity. Persistent phosphorylation of Thr588 (activation loop) promoted ubiquitinylation and proteasome-mediated degradation of DKF-1. Each DKF-1 domain displayed novel properties indicative of functional malleability (plasticity).",
author = "Hui Feng and Min Ren and Rubin, {Charles S.}",
year = "2006",
month = "6",
day = "30",
doi = "10.1074/jbc.M511898200",
language = "English (US)",
volume = "281",
pages = "17815--17826",
journal = "Journal of Biological Chemistry",
issn = "0021-9258",
publisher = "American Society for Biochemistry and Molecular Biology Inc.",
number = "26",

}

TY - JOUR

T1 - Conserved domains subserve novel mechanisms and functions in DKF-1, a Caenorhabditis elegans protein kinase D

AU - Feng, Hui

AU - Ren, Min

AU - Rubin, Charles S.

PY - 2006/6/30

Y1 - 2006/6/30

N2 - Protein kinase D (PKD) isoforms are effectors in signaling pathways controlled by diacylglycerol. PKDs contain conserved diacylglycerol binding (C1a, C1b), pleckstrin homology (PH), and Ser/Thr kinase domains. However, the properties of conserved domains may vary within the context of distinct PKD polypeptides. Such functional/structural malleability (plasticity) was explored by studying Caenorhabditis elegans D kinase family-1 (DKF-1), a PKD that governs locomotion in vivo. Phorbol ester binding with C1b alone activates classical PKDs by relieving C1-mediated inhibition. In contrast, C1a avidly ligated phorbol 12-myristate 13-acetate (PMA) and anchored DKF-1 at the plasma membrane. C1b bound PMA (moderate affinity) and cooperated with C1a in targeting DKF-1 to membranes. Mutations at a "Pro11" position in C1 domains were inactivating; kinase activity was minimal at PMA concentrations that stimulated wild type DKF-1 ∼10-fold. DKF-1 mutants exhibited unchanged, maximum kinase activity after cells were incubated with high PMA concentrations. Titration in situ revealed that translocation and activation of wild type and mutant DKF-1 were tightly and quantitatively linked at all PMA concentrations. Thus, C1 domains positively regulated phosphotransferase activity by docking DKF-1 with pools of activating lipid. A PH domain inhibits kinase activity in classical PKDs. The DKF-1 PH module neither inhibited catalytic activity nor bound phosphoinositides. Consequently, the PH module is an obligatory, positive regulator of DKF-1 activity that is compromised by mutation of Lys298 or Trp396. Phosphorylation of Thr588 switched on DKF-1 kinase activity. Persistent phosphorylation of Thr588 (activation loop) promoted ubiquitinylation and proteasome-mediated degradation of DKF-1. Each DKF-1 domain displayed novel properties indicative of functional malleability (plasticity).

AB - Protein kinase D (PKD) isoforms are effectors in signaling pathways controlled by diacylglycerol. PKDs contain conserved diacylglycerol binding (C1a, C1b), pleckstrin homology (PH), and Ser/Thr kinase domains. However, the properties of conserved domains may vary within the context of distinct PKD polypeptides. Such functional/structural malleability (plasticity) was explored by studying Caenorhabditis elegans D kinase family-1 (DKF-1), a PKD that governs locomotion in vivo. Phorbol ester binding with C1b alone activates classical PKDs by relieving C1-mediated inhibition. In contrast, C1a avidly ligated phorbol 12-myristate 13-acetate (PMA) and anchored DKF-1 at the plasma membrane. C1b bound PMA (moderate affinity) and cooperated with C1a in targeting DKF-1 to membranes. Mutations at a "Pro11" position in C1 domains were inactivating; kinase activity was minimal at PMA concentrations that stimulated wild type DKF-1 ∼10-fold. DKF-1 mutants exhibited unchanged, maximum kinase activity after cells were incubated with high PMA concentrations. Titration in situ revealed that translocation and activation of wild type and mutant DKF-1 were tightly and quantitatively linked at all PMA concentrations. Thus, C1 domains positively regulated phosphotransferase activity by docking DKF-1 with pools of activating lipid. A PH domain inhibits kinase activity in classical PKDs. The DKF-1 PH module neither inhibited catalytic activity nor bound phosphoinositides. Consequently, the PH module is an obligatory, positive regulator of DKF-1 activity that is compromised by mutation of Lys298 or Trp396. Phosphorylation of Thr588 switched on DKF-1 kinase activity. Persistent phosphorylation of Thr588 (activation loop) promoted ubiquitinylation and proteasome-mediated degradation of DKF-1. Each DKF-1 domain displayed novel properties indicative of functional malleability (plasticity).

UR - http://www.scopus.com/inward/record.url?scp=33745838566&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=33745838566&partnerID=8YFLogxK

U2 - 10.1074/jbc.M511898200

DO - 10.1074/jbc.M511898200

M3 - Article

C2 - 16613842

AN - SCOPUS:33745838566

VL - 281

SP - 17815

EP - 17826

JO - Journal of Biological Chemistry

JF - Journal of Biological Chemistry

SN - 0021-9258

IS - 26

ER -