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PMID: 7565670 Published · ppublish English Journal Article Research Support, Non-U.S. Gov't Research Support, U.S. Gov't, P.H.S.

A proline-rich sequence unique to MEK1 and MEK2 is required for raf binding and regulates MEK function.

Molecular and cellular biology ·Vol. 15 ·No. 10 ·1995-10-00 ·Pages 5214-25

Catling AD, Schaeffer HJ, Reuter CW, Reddy GR, Weber MJ

Abstract

Mammalian MEK1 and MEK2 contain a proline-rich (PR) sequence that is absent both from the yeast homologs Ste7 and Byr1 and from a recently cloned activator of the JNK/stress-activated protein kinases, SEK1/MKK4. Since this PR sequence occurs in MEKs that are regulated by Raf family enzymes but is missing from MEKs and SEKs activated independently of Raf, we sought to investigate the role of this sequence in MEK1 and MEK2 regulation and function. Deletion of the PR sequence from MEK1 blocked the ability of MEK1 to associate with members of the Raf family and markedly attenuated activation of the protein in vivo following growth factor stimulation. In addition, this sequence was necessary for efficient activation of MEK1 in vitro by B-Raf but dispensable for activation by a novel MEK1 activator which we have previously detected in fractionated fibroblast extracts. Furthermore, we found that a phosphorylation site within the PR sequence of MEK1 was required for sustained MEK1 activity in response to serum stimulation of quiescent fibroblasts. Consistent with this observation, we observed that MEK2, which lacks a phosphorylation site at the corresponding position, was activated only transiently following serum stimulation. Finally, we found that deletion of the PR sequence from a constitutively activated MEK1 mutant rendered the protein nontransforming in Rat1 fibroblasts. These observations indicate a critical role for the PR sequence in directing specific protein-protein interactions important for the activation, inactivation, and downstream functioning of the MEKs.

MeSH Terms
Amino Acid Sequence Animals Blood Cell Line Cell Transformation, Neoplastic Enzyme Activation Fibroblasts MAP Kinase Kinase 1 MAP Kinase Kinase 2 Mitogen-Activated Protein Kinase Kinases Molecular Sequence Data Peptide Mapping Phosphorylation Proline/physiology Protein Serine-Threonine Kinases/genetics,metabolism,physiology Protein-Tyrosine Kinases/genetics,metabolism,physiology Proto-Oncogene Proteins/metabolism Proto-Oncogene Proteins c-raf Rats Recombinant Fusion Proteins/biosynthesis Sequence Deletion Threonine/metabolism
Chemicals
Proto-Oncogene Proteins Recombinant Fusion Proteins Threonine Proline Protein-Tyrosine Kinases Protein Serine-Threonine Kinases Proto-Oncogene Proteins c-raf MAP Kinase Kinase 1 MAP Kinase Kinase 2 Mitogen-Activated Protein Kinase Kinases
Authors & Affiliations
5 authors, click to expand affiliations / ORCID
Catling A D
Department of Microbiology and Cancer Center, University of Virginia Health Sciences Center, Charlottesville 22908, USA.
Schaeffer H J
Reuter C W
Reddy G R
Weber M J
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Article Info
Journal
Molecular and cellular biology
Abbr.
Mol Cell Biol
ISSN
0270-7306
DOI
10.1128/MCB.15.10.5214
Published
1995-10-00
Pages
5214-25
Language
English
Region
United States
NLM ID
8109087
PMCID
PMC230769
Subset
IM
Grants
NIGMS NIH HHS · GM47332 · United States
External Links
PubMed source DOI: 10.1128/MCB.15.10.5214 PMC Full Text View journal details
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