Signal Transduction
Functional Cross-talk between Ras and Rho Pathways: A Ras-SPECIFIC GTPase-ACTIVATING PROTEIN (p120RasGAP) COMPETITIVELY INHIBITS THE RhoGAP ACTIVITY OF DELETED IN LIVER CANCER (DLC) TUMOR SUPPRESSOR BY MASKING THE CATALYTIC ARGININE FINGER*

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The three deleted in liver cancer genes (DLC1–3) encode Rho-specific GTPase-activating proteins (RhoGAPs). Their expression is frequently silenced in a variety of cancers. The RhoGAP activity, which is required for full DLC-dependent tumor suppressor activity, can be inhibited by the Src homology 3 (SH3) domain of a Ras-specific GAP (p120RasGAP). Here, we comprehensively investigated the molecular mechanism underlying cross-talk between two distinct regulators of small GTP-binding proteins using structural and biochemical methods. We demonstrate that only the SH3 domain of p120 selectively inhibits the RhoGAP activity of all three DLC isoforms as compared with a large set of other representative SH3 or RhoGAP proteins. Structural and mutational analyses provide new insights into a putative interaction mode of the p120 SH3 domain with the DLC1 RhoGAP domain that is atypical and does not follow the classical PXXP-directed interaction. Hence, p120 associates with the DLC1 RhoGAP domain by targeting the catalytic arginine finger and thus by competitively and very potently inhibiting RhoGAP activity. The novel findings of this study shed light on the molecular mechanisms underlying the DLC inhibitory effects of p120 and suggest a functional cross-talk between Ras and Rho proteins at the level of regulatory proteins.

GTPase
Kinetics
Molecular Modeling
SH3 Domains
Structural Biology
Arginine Finger
DLC1
Deleted in Liver Cancer
RhoGAP
p120RasGAP

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*

This work was supported in part by the German Research Foundation (Deutsche Forschungsgemeinschaft (DFG)) through the Collaborative Research Center 974 (SFB 974) “Communication and Systems Relevance during Liver Injury and Regeneration,” the International Research Training Group 1902 (IRGT1902), and Project Grant AH 92/5-1; the National Genome Research Network Plus program of the German Ministry of Science and Education (Bundesministerium für Bildung und Forschung Grant 01GS08100); and the International North Rhine-Westphalia Research School BioStruct granted by the Ministry of Innovation, Science and Research of the State North Rhine-Westphalia, the Heinrich Heine University of Düsseldorf, and the Entrepreneur Foundation at the Heinrich Heine University of Düsseldorf.

6

M. Jaiswal, E. Amin, and R. Dvorsky, unpublished data.

1

Present address: Structural Biology Group, Max Planck Inst. for Molecular Physiology, Otto-Hahn-Strasse 11, 44227 Dortmund, Germany.

2

Present address: Inst. of Physical Biology, Heinrich Heine University, 40255 Düsseldorf, Germany.

3

Supported by the DFG Heisenberg program.

5

The abbreviations used are:

    GAP

    GTPase-activating protein

    DLC

    deleted in liver cancer

    SH

    Src homology

    SAM

    sterile α motif

    START

    steroidogenic acute regulatory related lipid transfer

    aa

    amino acids

    tamra

    tetramethylrhodamine

    aSEC

    analytical size exclusion chromatography

    ITC

    isothermal titration calorimetry.