ARF-GEF cytohesin-2/ARNO regulates R-Ras and α5-integrin recycling through an EHD1-positive compartment

Joseph C. Salem, Marta M. Reviriego-Mendoza, Lorraine C. Santy

Research output: Contribution to journalArticle

4 Citations (Scopus)

Abstract

When expressed in epithelial cells, cytohesin-2/ARNO, a guanine nucleotide exchange factor (GEF) for ARF small GTPases, causes a robust migration response. Recent evidence suggests that cytohesin-2/ARNO acts downstream of small the GTPase R-Ras to promote spreading and migration. We hypothesized that cytohesin-2/ARNO could transmit R-Ras signals by regulating the recycling of R-Ras through ARF activation. We found that Eps15-homology domain 1(EHD1), a protein that associates with the endocytic recycling compartment (ERC), colocalizes with active R-Ras in transiently expressed HeLa cells. In addition, we show that EHD1-positive recycling endosomes are a novel compartment for cytohesin-2/ARNO. Knockdown or expression of GEF-inactive (E156K) cytohesin-2/ARNO causes R-Ras to accumulate on recycling endosomes containing EHD1 and inhibits cell spreading. E156K-ARNO also causes a reduction in focal adhesion size and number. Finally, we demonstrate that R-Ras/ARNO signaling is required for recycling of α5-integrin and R-Ras to the plasma membrane. These data establish a role for cytohesin-2/ARNO as a regulator of R-Ras and integrin recycling and suggest that ARF-regulated trafficking of R-Ras is required for R-Ras-dependent effects on spreading and adhesion formation.

Original languageEnglish (US)
Pages (from-to)4265-4279
Number of pages15
JournalMolecular biology of the cell
Volume26
Issue number23
DOIs
StatePublished - Nov 15 2015

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Guanine Nucleotide Exchange Factors
Recycling
Integrins
Monomeric GTP-Binding Proteins
Endosomes
Focal Adhesions
HeLa Cells
cytohesin-2
Epithelial Cells
Cell Membrane
Proteins

All Science Journal Classification (ASJC) codes

  • Molecular Biology
  • Cell Biology

Cite this

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title = "ARF-GEF cytohesin-2/ARNO regulates R-Ras and α5-integrin recycling through an EHD1-positive compartment",
abstract = "When expressed in epithelial cells, cytohesin-2/ARNO, a guanine nucleotide exchange factor (GEF) for ARF small GTPases, causes a robust migration response. Recent evidence suggests that cytohesin-2/ARNO acts downstream of small the GTPase R-Ras to promote spreading and migration. We hypothesized that cytohesin-2/ARNO could transmit R-Ras signals by regulating the recycling of R-Ras through ARF activation. We found that Eps15-homology domain 1(EHD1), a protein that associates with the endocytic recycling compartment (ERC), colocalizes with active R-Ras in transiently expressed HeLa cells. In addition, we show that EHD1-positive recycling endosomes are a novel compartment for cytohesin-2/ARNO. Knockdown or expression of GEF-inactive (E156K) cytohesin-2/ARNO causes R-Ras to accumulate on recycling endosomes containing EHD1 and inhibits cell spreading. E156K-ARNO also causes a reduction in focal adhesion size and number. Finally, we demonstrate that R-Ras/ARNO signaling is required for recycling of α5-integrin and R-Ras to the plasma membrane. These data establish a role for cytohesin-2/ARNO as a regulator of R-Ras and integrin recycling and suggest that ARF-regulated trafficking of R-Ras is required for R-Ras-dependent effects on spreading and adhesion formation.",
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ARF-GEF cytohesin-2/ARNO regulates R-Ras and α5-integrin recycling through an EHD1-positive compartment. / Salem, Joseph C.; Reviriego-Mendoza, Marta M.; Santy, Lorraine C.

In: Molecular biology of the cell, Vol. 26, No. 23, 15.11.2015, p. 4265-4279.

Research output: Contribution to journalArticle

TY - JOUR

T1 - ARF-GEF cytohesin-2/ARNO regulates R-Ras and α5-integrin recycling through an EHD1-positive compartment

AU - Salem, Joseph C.

AU - Reviriego-Mendoza, Marta M.

AU - Santy, Lorraine C.

PY - 2015/11/15

Y1 - 2015/11/15

N2 - When expressed in epithelial cells, cytohesin-2/ARNO, a guanine nucleotide exchange factor (GEF) for ARF small GTPases, causes a robust migration response. Recent evidence suggests that cytohesin-2/ARNO acts downstream of small the GTPase R-Ras to promote spreading and migration. We hypothesized that cytohesin-2/ARNO could transmit R-Ras signals by regulating the recycling of R-Ras through ARF activation. We found that Eps15-homology domain 1(EHD1), a protein that associates with the endocytic recycling compartment (ERC), colocalizes with active R-Ras in transiently expressed HeLa cells. In addition, we show that EHD1-positive recycling endosomes are a novel compartment for cytohesin-2/ARNO. Knockdown or expression of GEF-inactive (E156K) cytohesin-2/ARNO causes R-Ras to accumulate on recycling endosomes containing EHD1 and inhibits cell spreading. E156K-ARNO also causes a reduction in focal adhesion size and number. Finally, we demonstrate that R-Ras/ARNO signaling is required for recycling of α5-integrin and R-Ras to the plasma membrane. These data establish a role for cytohesin-2/ARNO as a regulator of R-Ras and integrin recycling and suggest that ARF-regulated trafficking of R-Ras is required for R-Ras-dependent effects on spreading and adhesion formation.

AB - When expressed in epithelial cells, cytohesin-2/ARNO, a guanine nucleotide exchange factor (GEF) for ARF small GTPases, causes a robust migration response. Recent evidence suggests that cytohesin-2/ARNO acts downstream of small the GTPase R-Ras to promote spreading and migration. We hypothesized that cytohesin-2/ARNO could transmit R-Ras signals by regulating the recycling of R-Ras through ARF activation. We found that Eps15-homology domain 1(EHD1), a protein that associates with the endocytic recycling compartment (ERC), colocalizes with active R-Ras in transiently expressed HeLa cells. In addition, we show that EHD1-positive recycling endosomes are a novel compartment for cytohesin-2/ARNO. Knockdown or expression of GEF-inactive (E156K) cytohesin-2/ARNO causes R-Ras to accumulate on recycling endosomes containing EHD1 and inhibits cell spreading. E156K-ARNO also causes a reduction in focal adhesion size and number. Finally, we demonstrate that R-Ras/ARNO signaling is required for recycling of α5-integrin and R-Ras to the plasma membrane. These data establish a role for cytohesin-2/ARNO as a regulator of R-Ras and integrin recycling and suggest that ARF-regulated trafficking of R-Ras is required for R-Ras-dependent effects on spreading and adhesion formation.

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