Using EPAC1 (exchange protein directly activated by cAMP 1) as bait in two-hybrid screens of foetal and adult human brain libraries, we identified the LC2 (light chain 2) of MAP1A (microtubule-associated protein 1A) as a protein capable of interaction with EPAC1. We applied an immunoprecipitation assay to demonstrate protein interaction between EPAC1 and LC2 in co-transfected human embryonic kidney 293 cells. EPAC2 also co-immunoprecipitated with LC2 from extracts of rat cerebellum. Immunolocalization in co-transfected human embryonic kidney 293 cells revealed that EPAC1 co-localizes with LC2 throughout the cell body. We found that endogenous EPAC2 is also immunolocalized with LC2 in PC12 cells. Immunolocalization of EPAC1 in transfected COS1 cells showed that EPAC1 is associated with the perinuclear region surrounding the nucleus and filamentous structures throughout the cell. Removal of the cAMP-binding domain of EPAC1 (ΔcAMP-EPAC1) appeared to disrupt targeting of EPAC1 in cells resulting in a more dispersed staining pattern. Using two-hybrid assay, we tested the ability of LC2 to interact with ΔcAMP-EPAC1 and ΔDEP-EPAC1, which lacks a DEP domain (dishevelled, Egl-10 and pleckstrin homology domain). We found that deletion of the cAMP-binding domain inhibited interaction between EPAC1 and LC2 in a two-hybrid assay, but removal of the DEP domain had little effect. LC2 was found to interact with a glutathione-S-transferase-fusion protein of the cAMP-binding domain of EPAC1 in a pull-down assay, but not the DEP, REM (Ras exchange motif) or CAT (catalytic) domains. Together with our two-hybrid results, this suggests that the cAMP-binding domain of EPAC1 mediates interaction with LC2.
Skip Nav Destination
Follow us on Twitter @Biochem_Journal
Article navigation
September 2004
-
Cover Image
Cover Image
- PDF Icon PDF LinkFront Matter
- PDF Icon PDF LinkTable of Contents
- PDF Icon PDF LinkEditorial Board
Research Article|
September 07 2004
Exchange protein directly activated by cAMP (EPAC) interacts with the light chain (LC) 2 of MAP1A
Maria M. MAGIERA;
Maria M. MAGIERA
1Molecular Pharmacology Group, Division of Biochemistry and Molecular Biology, Faculty of Biomedical and Life Sciences, University of Glasgow, Davidson Building, Glasgow G12 8QQ, Scotland, U.K.
Search for other works by this author on:
Mona GUPTA;
Mona GUPTA
1Molecular Pharmacology Group, Division of Biochemistry and Molecular Biology, Faculty of Biomedical and Life Sciences, University of Glasgow, Davidson Building, Glasgow G12 8QQ, Scotland, U.K.
Search for other works by this author on:
Catherine J. RUNDELL;
Catherine J. RUNDELL
1Molecular Pharmacology Group, Division of Biochemistry and Molecular Biology, Faculty of Biomedical and Life Sciences, University of Glasgow, Davidson Building, Glasgow G12 8QQ, Scotland, U.K.
Search for other works by this author on:
Nilima SATISH;
Nilima SATISH
1Molecular Pharmacology Group, Division of Biochemistry and Molecular Biology, Faculty of Biomedical and Life Sciences, University of Glasgow, Davidson Building, Glasgow G12 8QQ, Scotland, U.K.
Search for other works by this author on:
Isabelle ERNENS;
Isabelle ERNENS
1Molecular Pharmacology Group, Division of Biochemistry and Molecular Biology, Faculty of Biomedical and Life Sciences, University of Glasgow, Davidson Building, Glasgow G12 8QQ, Scotland, U.K.
Search for other works by this author on:
Stephen J. YARWOOD
Stephen J. YARWOOD
1
1Molecular Pharmacology Group, Division of Biochemistry and Molecular Biology, Faculty of Biomedical and Life Sciences, University of Glasgow, Davidson Building, Glasgow G12 8QQ, Scotland, U.K.
1To whom correspondence should be addressed (email [email protected]).
Search for other works by this author on:
Publisher: Portland Press Ltd
Received:
January 21 2004
Revision Received:
May 13 2004
Accepted:
June 18 2004
Accepted Manuscript online:
June 18 2004
Online ISSN: 1470-8728
Print ISSN: 0264-6021
The Biochemical Society, London
2004
Biochem J (2004) 382 (3): 803–810.
Article history
Received:
January 21 2004
Revision Received:
May 13 2004
Accepted:
June 18 2004
Accepted Manuscript online:
June 18 2004
Citation
Maria M. MAGIERA, Mona GUPTA, Catherine J. RUNDELL, Nilima SATISH, Isabelle ERNENS, Stephen J. YARWOOD; Exchange protein directly activated by cAMP (EPAC) interacts with the light chain (LC) 2 of MAP1A. Biochem J 15 September 2004; 382 (3): 803–810. doi: https://doi.org/10.1042/BJ20040122
Download citation file:
Sign in
Don't already have an account? Register
Sign in to your personal account
You could not be signed in. Please check your email address / username and password and try again.
Could not validate captcha. Please try again.
Biochemical Society Member Sign in
Sign InSign in via your Institution
Sign in via your InstitutionGet Access To This Article
Follow us on Twitter @Biochem_Journal
Open Access for all
We offer compliant routes for all authors from 2025. With library support, there will be no author nor reader charges in 5 journals. Check here |
![]() View past webinars > |