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Cited by 679Protein Kinase Regulation and GTPase SignalingCellular transport and secretionCellular Mechanics and Interactions

Differential Localization of Rho Gtpases in Live Cells

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Michaelson, David, Silletti, Joseph, Murphy, Gretchen, D'Eustachio, Peter, Rush, Mark, Philips, Mark R.·2001·The Journal of Cell Biology
DOI 10.1083/jcb.152.1.111PMC PMC2193662
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Abstract

Determinants of membrane targeting of Rho proteins were investigated in live cells with green fluorescent fusion proteins expressed with or without Rho-guanine nucleotide dissociation inhibitor (GDI)α. The hypervariable region determined to which membrane compartment each protein was targeted. Targeting was regulated by binding to RhoGDIα in the case of RhoA, Rac1, Rac2, and Cdc42hs but not RhoB or TC10. Although RhoB localized to the plasma membrane (PM), Golgi, and motile peri-Golgi vesicles, TC10 localized to PMs and endosomes. Inhibition of palmitoylation mislocalized H-Ras, RhoB, and TC10 to the endoplasmic reticulum. Although overexpressed Cdc42hs and Rac2 were observed predominantly on endomembrane, Rac1 was predominantly at the PM. RhoA was cytosolic even when expressed at levels in vast excess of RhoGDIα. Oncogenic Dbl stimulated translocation of green fluorescent protein (GFP)-Rac1, GFP-Cdc42hs, and GFP-RhoA to lamellipodia. RhoGDI binding to GFP-Cdc42hs was not affected by substituting farnesylation for geranylgeranylation. A palmitoylation site inserted into RhoA blocked RhoGDIα binding. Mutations that render RhoA, Cdc42hs, or Rac1, either constitutively active or dominant negative abrogated binding to RhoGDIα and redirected expression to both PMs and internal membranes. Thus, despite the common essential feature of the CAAX (prenylation, AAX tripeptide proteolysis, and carboxyl methylation) motif, the subcellular localizations of Rho GTPases, like their functions, are diverse and dynamic.

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  • Active Rho GTPases are localized on the plasma membrane.

    “The difference between wild-type Rho GTPases and constitutively active mutants presumably reflects the difference in the fraction of the protein bound to the plasma membrane, as active Rho GTPases are localized on the plasma membrane21.”

    From Local, persistent activation of Rho GTPases during plasticity of single dendritic spines (2011) · cites this paper as [21]

    The Discussion (page unavailable) reports that Dbl-stimulated Rho GTPases associated predominantly with plasma-membrane regions. But it also states that constitutively active mutants localized to both the plasma membrane and internal membranes, so the claim overstates plasma-membrane localization as a general rule for active Rho GTPases. (LLM, conf=0.88)

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Authors
Michaelson, D., Silletti, J., Murphy, G., D'Eustachio, P., Rush, M., & Philips, M. R.
Affiliations
  • New York University (US)
  • cDepartment of Biochemistry, New York University School of Medicine, New York, New York 10016
ORCID
Philips, Mark R.
Venue
The Journal of Cell Biology
Publisher
Rockefeller University Press
Published
2001-01-08
Volume
152
Issue
1
Pages
111-126
ISSN
  • 0021-9525 (ISSN-L)
  • 1540-8140
DOI
10.1083/jcb.152.1.111
PMC
PMC2193662
OpenAlex
W2165803049
Cited by
679
References
0
Citations by year
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FWCI
14.24
Type
journal article
Language
en
Open access
Licence
cc-by-nc-sa
Locations
  • The Journal of Cell Biology (publishedVersion)
  • PubMed Central (publishedVersion, cc-by-nc-sa)
  • PubMed (publishedVersion)
Funders
  • National Institute of General Medical Sciences (GM55279, R01GM055279, T32GM007308)
  • National Institute of Allergy and Infectious Diseases (R01AI036224, R29AI036224, AI-36224)
  • National Center for Research Resources (M01RR000096)
Topics
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Protein Kinase Regulation and GTPase Signaling
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Cell Biology › Cellular transport and secretion
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Cell Biology › Cellular Mechanics and Interactions
Keywords
Rho GTPases, membrane targeting, subcellular localization, palmitoylation, farnesylation, CAAX motif, RhoA, Rac1, Rac2, Cdc42, RhoB, TC10, H-Ras, Dbl, lamellipodia, plasma membrane, Golgi apparatus, endosomes, endoplasmic reticulum, guanine nucleotide dissociation inhibitor
MeSH terms
Amino Acid Sequence, Animals, Biological Transport, Cell Line, Cell Membrane, Chlorocebus aethiops, Dogs, Cricetinae, Luminescent Proteins, Molecular Sequence Data, Recombinant Fusion Proteins, CHO Cells, 3T3 Cells, Palmitic Acid, COS Cells, Guanine Nucleotide Dissociation Inhibitors, rho GTP-Binding Proteins, Green Fluorescent Proteins, Mice, rho-Specific Guanine Nucleotide Dissociation Inhibitors

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