DOI: 10.1091/mbc.10.6.1851. Activation of RhoA by Lysophosphatidic Acid and Gα<sub>12/13</sub>Subunits in Neuronal Cells: Induction of Neurite Retraction

Activation of RhoA by Lysophosphatidic Acid and Gα12/13Subunits in Neuronal Cells: Induction of Neurite Retraction

10.1091/mbc.10.6.1851

Crossref journal-article

American Society for Cell Biology (ASCB)

Molecular Biology of the Cell (1076)

Abstract

Neuronal cells undergo rapid growth cone collapse, neurite retraction, and cell rounding in response to certain G protein–coupled receptor agonists such as lysophosphatidic acid (LPA). These shape changes are driven by Rho-mediated contraction of the actomyosin-based cytoskeleton. To date, however, detection of Rho activation has been hampered by the lack of a suitable assay. Furthermore, the nature of the G protein(s) mediating LPA-induced neurite retraction remains unknown. We have developed a Rho activation assay that is based on the specific binding of active RhoA to its downstream effector Rho-kinase (ROK). A fusion protein of GST and the Rho-binding domain of ROK pulls down activated but not inactive RhoA from cell lysates. Using GST-ROK, we show that in N1E-115 neuronal cells LPA activates endogenous RhoA within 30 s, concomitant with growth cone collapse. Maximal activation occurs after 3 min when neurite retraction is complete and the actin cytoskeleton is fully contracted. LPA-induced RhoA activation is completely inhibited by tyrosine kinase inhibitors (tyrphostin 47 and genistein). Activated Gα12and Gα13subunits mimic LPA both in activating RhoA and in inducing RhoA-mediated cytoskeletal contraction, thereby preventing neurite outgrowth. We conclude that in neuronal cells, LPA activates RhoA to induce growth cone collapse and neurite retraction through a G12/13-initiated pathway that involves protein-tyrosine kinase activity.

Bibliography

Kranenburg, O., Poland, M., van Horck, F. P. G., Drechsel, D., Hall, A., & Moolenaar, W. H. (1999). Activation of RhoA by Lysophosphatidic Acid and GÎ±12/13Subunits in Neuronal Cells: Induction of Neurite Retraction. Molecular Biology of the Cell, 10(6), 1851â1857.

Authors 6

Onno Kranenburg (first)
Mieke Poland (additional)
Francis P. G. van Horck (additional)
David Drechsel (additional)
Alan Hall (additional)
Wouter H. Moolenaar (additional)

References 33 Referenced 256

10.1126/science.275.5304.1308 / Science by Amano M. (1997)
10.1046/j.1365-2443.1998.00181.x / Genes Cells by Amano M. (1998)
10.1074/jbc.270.42.24631 / J. Biol. Chem. by Buhl A.M. (1995)
10.1038/sj.onc.1201005 / Oncogene by de Rooij J. (1997)
10.1093/emboj/16.2.252 / EMBO J. by Franke B. (1997)
10.1073/pnas.94.19.10098 / Proc. Natl. Acad. Sci. USA by Fromm C. (1997)
10.1083/jcb.137.7.1603 / J. Cell Biol. by Gebbink M.F. (1997)
10.1074/jbc.273.8.4653 / J. Biol. Chem. by Gohla A. (1998)
10.1074/jbc.273.4.1839 / J. Biol. Chem. by Gutkind J.S. (1998)
10.1126/science.279.5350.509 / Science by Hall A. (1998)
10.1126/science.280.5372.2112 / Science by Hart M.J. (1998)
10.1128/MCB.16.2.648 / Mol. Cell. Biol. by Heasley L.E. (1996)
10.1083/jcb.141.7.1625 / J. Cell Biol. by Hirose M. (1998)
{'key': 'B14', 'first-page': '247', 'volume': '4', 'author': 'Jalink K.', 'year': '1993', 'journal-title': 'Cell Growth & Differ.'} / Cell Growth & Differ. by Jalink K. (1993)
10.1083/jcb.118.2.411 / J. Cell Biol. by Jalink K. (1992)
10.1083/jcb.126.3.801 / J. Cell Biol. by Jalink K. (1994)
10.1074/jbc.273.5.2489 / J. Biol. Chem. by Katoh H. (1998)
10.1074/jbc.273.44.28700 / J. Biol. Chem. by Katoh H. (1998)
10.1128/MCB.17.3.1201 / Mol. Cell. Biol. by Kozma R. (1997)
10.1242/jcs.110.19.2417 / J. Cell Sci. by Kranenburg O. (1997)
10.1016/S0092-8674(00)81371-9 / Cell by Lamarche N. (1996)
10.1128/MCB.16.10.5313 / Mol. Cell. Biol. by Leung T. (1996)
10.1074/jbc.270.49.29051 / J. Biol. Chem. by Leung T. (1995)
10.1016/S1097-2765(00)80019-2 / Mol. Cell by Manser E. (1998)
10.1093/emboj/17.19.5638 / EMBO J. by Mao J. (1998)
10.1002/j.1460-2075.1996.tb00574.x / EMBO J. by Matsui T. (1996)
10.1016/S0955-0674(97)80059-2 / Curr. Opin. Cell Biol. by Moolenaar W.H. (1997)
10.1242/jcs.108.1.225 / J. Cell Sci. by Nobes C.D. (1995)
10.1016/0092-8674(92)90163-7 / Cell by Ridley A.J. (1992)
10.1016/S0960-9822(02)70785-9 / Curr. Biol. by Taylor S.J. (1996)
10.1101/gad.11.18.2295 / Genes & Dev. by Van Aelst L. (1997)
10.1083/jcb.139.3.797 / J. Cell Biol. by van Leeuwen F.N. (1997)
10.1073/pnas.90.14.6741 / Proc. Natl. Acad. Sci. USA by Xu N. (1993)

Dates

Type	When
Created	12 years, 1 month ago (July 2, 2013, 7:32 p.m.)
Deposited	3 years, 5 months ago (Feb. 26, 2022, 9:39 p.m.)
Indexed	1 month ago (July 19, 2025, 11:26 p.m.)
Issued	26 years, 2 months ago (June 1, 1999)
Published	26 years, 2 months ago (June 1, 1999)
Published Print	26 years, 2 months ago (June 1, 1999)

Funders 0

None

BibTeX

@article{Kranenburg_1999, title={Activation of RhoA by Lysophosphatidic Acid and Gα12/13Subunits in Neuronal Cells: Induction of Neurite Retraction}, volume={10}, ISSN={1939-4586}, url={http://dx.doi.org/10.1091/mbc.10.6.1851}, DOI={10.1091/mbc.10.6.1851}, number={6}, journal={Molecular Biology of the Cell}, publisher={American Society for Cell Biology (ASCB)}, author={Kranenburg, Onno and Poland, Mieke and van Horck, Francis P. G. and Drechsel, David and Hall, Alan and Moolenaar, Wouter H.}, editor={Beckerle, Mary C.}, year={1999}, month=jun, pages={1851–1857} }

JSON

{
  "indexed": {
    "date-parts": [
      [
        2025,
        7,
        20
      ]
    ],
    "date-time": "2025-07-20T03:26:55Z",
    "timestamp": 1752982015821
  },
  "reference-count": 33,
  "publisher": "American Society for Cell Biology (ASCB)",
  "issue": "6",
  "content-domain": {
    "domain": [],
    "crossmark-restriction": false
  },
  "published-print": {
    "date-parts": [
      [
        1999,
        6
      ]
    ]
  },
  "abstract": "<jats:p>Neuronal cells undergo rapid growth cone collapse, neurite retraction, and cell rounding in response to certain G protein\u2013coupled receptor agonists such as lysophosphatidic acid (LPA). These shape changes are driven by Rho-mediated contraction of the actomyosin-based cytoskeleton. To date, however, detection of Rho activation has been hampered by the lack of a suitable assay. Furthermore, the nature of the G protein(s) mediating LPA-induced neurite retraction remains unknown. We have developed a Rho activation assay that is based on the specific binding of active RhoA to its downstream effector Rho-kinase (ROK). A fusion protein of GST and the Rho-binding domain of ROK pulls down activated but not inactive RhoA from cell lysates. Using GST-ROK, we show that in N1E-115 neuronal cells LPA activates endogenous RhoA within 30 s, concomitant with growth cone collapse. Maximal activation occurs after 3 min when neurite retraction is complete and the actin cytoskeleton is fully contracted. LPA-induced RhoA activation is completely inhibited by tyrosine kinase inhibitors (tyrphostin 47 and genistein). Activated G\u03b1<jats:sub>12</jats:sub>and G\u03b1<jats:sub>13</jats:sub>subunits mimic LPA both in activating RhoA and in inducing RhoA-mediated cytoskeletal contraction, thereby preventing neurite outgrowth. We conclude that in neuronal cells, LPA activates RhoA to induce growth cone collapse and neurite retraction through a G<jats:sub>12/13</jats:sub>-initiated pathway that involves protein-tyrosine kinase activity.</jats:p>",
  "DOI": "10.1091/mbc.10.6.1851",
  "type": "journal-article",
  "created": {
    "date-parts": [
      [
        2013,
        7,
        2
      ]
    ],
    "date-time": "2013-07-02T23:32:15Z",
    "timestamp": 1372807935000
  },
  "page": "1851-1857",
  "source": "Crossref",
  "is-referenced-by-count": 256,
  "title": "Activation of RhoA by Lysophosphatidic Acid and G\u03b1<sub>12/13</sub>Subunits in Neuronal Cells: Induction of Neurite Retraction",
  "prefix": "10.1091",
  "volume": "10",
  "author": [
    {
      "given": "Onno",
      "family": "Kranenburg",
      "sequence": "first",
      "affiliation": [
        {
          "name": "The Netherlands Cancer Institute, Division of Cellular Biochemistry, 1066 CX Amsterdam, The Netherlands; and"
        }
      ]
    },
    {
      "given": "Mieke",
      "family": "Poland",
      "sequence": "additional",
      "affiliation": [
        {
          "name": "The Netherlands Cancer Institute, Division of Cellular Biochemistry, 1066 CX Amsterdam, The Netherlands; and"
        }
      ]
    },
    {
      "given": "Francis P. G.",
      "family": "van Horck",
      "sequence": "additional",
      "affiliation": [
        {
          "name": "The Netherlands Cancer Institute, Division of Cellular Biochemistry, 1066 CX Amsterdam, The Netherlands; and"
        }
      ]
    },
    {
      "given": "David",
      "family": "Drechsel",
      "sequence": "additional",
      "affiliation": [
        {
          "name": "Medical Research Council Laboratory for Molecular Cell Biology, University College London, London WC1E 6BT, United Kingdom"
        }
      ]
    },
    {
      "given": "Alan",
      "family": "Hall",
      "sequence": "additional",
      "affiliation": [
        {
          "name": "Medical Research Council Laboratory for Molecular Cell Biology, University College London, London WC1E 6BT, United Kingdom"
        }
      ]
    },
    {
      "given": "Wouter H.",
      "family": "Moolenaar",
      "sequence": "additional",
      "affiliation": [
        {
          "name": "The Netherlands Cancer Institute, Division of Cellular Biochemistry, 1066 CX Amsterdam, The Netherlands; and"
        }
      ]
    }
  ],
  "member": "1076",
  "reference": [
    {
      "key": "B1",
      "doi-asserted-by": "crossref",
      "first-page": "1308",
      "DOI": "10.1126/science.275.5304.1308",
      "volume": "275",
      "author": "Amano M.",
      "year": "1997",
      "journal-title": "Science"
    },
    {
      "key": "B2",
      "doi-asserted-by": "crossref",
      "first-page": "177",
      "DOI": "10.1046/j.1365-2443.1998.00181.x",
      "volume": "3",
      "author": "Amano M.",
      "year": "1998",
      "journal-title": "Genes Cells"
    },
    {
      "key": "B3",
      "doi-asserted-by": "crossref",
      "first-page": "24631",
      "DOI": "10.1074/jbc.270.42.24631",
      "volume": "270",
      "author": "Buhl A.M.",
      "year": "1995",
      "journal-title": "J. Biol. Chem."
    },
    {
      "key": "B4",
      "doi-asserted-by": "crossref",
      "first-page": "623",
      "DOI": "10.1038/sj.onc.1201005",
      "volume": "14",
      "author": "de Rooij J.",
      "year": "1997",
      "journal-title": "Oncogene"
    },
    {
      "key": "B5",
      "doi-asserted-by": "crossref",
      "first-page": "252",
      "DOI": "10.1093/emboj/16.2.252",
      "volume": "16",
      "author": "Franke B.",
      "year": "1997",
      "journal-title": "EMBO J."
    },
    {
      "key": "B6",
      "doi-asserted-by": "crossref",
      "first-page": "10098",
      "DOI": "10.1073/pnas.94.19.10098",
      "volume": "94",
      "author": "Fromm C.",
      "year": "1997",
      "journal-title": "Proc. Natl. Acad. Sci. USA"
    },
    {
      "key": "B7",
      "doi-asserted-by": "crossref",
      "first-page": "1603",
      "DOI": "10.1083/jcb.137.7.1603",
      "volume": "137",
      "author": "Gebbink M.F.",
      "year": "1997",
      "journal-title": "J. Cell Biol."
    },
    {
      "key": "B8",
      "doi-asserted-by": "crossref",
      "first-page": "4653",
      "DOI": "10.1074/jbc.273.8.4653",
      "volume": "273",
      "author": "Gohla A.",
      "year": "1998",
      "journal-title": "J. Biol. Chem."
    },
    {
      "key": "B9",
      "doi-asserted-by": "crossref",
      "first-page": "1839",
      "DOI": "10.1074/jbc.273.4.1839",
      "volume": "273",
      "author": "Gutkind J.S.",
      "year": "1998",
      "journal-title": "J. Biol. Chem."
    },
    {
      "key": "B10",
      "doi-asserted-by": "crossref",
      "first-page": "509",
      "DOI": "10.1126/science.279.5350.509",
      "volume": "279",
      "author": "Hall A.",
      "year": "1998",
      "journal-title": "Science"
    },
    {
      "key": "B11",
      "doi-asserted-by": "crossref",
      "first-page": "2112",
      "DOI": "10.1126/science.280.5372.2112",
      "volume": "280",
      "author": "Hart M.J.",
      "year": "1998",
      "journal-title": "Science"
    },
    {
      "key": "B12",
      "doi-asserted-by": "crossref",
      "first-page": "648",
      "DOI": "10.1128/MCB.16.2.648",
      "volume": "16",
      "author": "Heasley L.E.",
      "year": "1996",
      "journal-title": "Mol. Cell. Biol."
    },
    {
      "key": "B13",
      "doi-asserted-by": "crossref",
      "first-page": "1625",
      "DOI": "10.1083/jcb.141.7.1625",
      "volume": "141",
      "author": "Hirose M.",
      "year": "1998",
      "journal-title": "J. Cell Biol."
    },
    {
      "key": "B14",
      "first-page": "247",
      "volume": "4",
      "author": "Jalink K.",
      "year": "1993",
      "journal-title": "Cell Growth & Differ."
    },
    {
      "key": "B15",
      "doi-asserted-by": "crossref",
      "first-page": "411",
      "DOI": "10.1083/jcb.118.2.411",
      "volume": "118",
      "author": "Jalink K.",
      "year": "1992",
      "journal-title": "J. Cell Biol."
    },
    {
      "key": "B16",
      "doi-asserted-by": "crossref",
      "first-page": "801",
      "DOI": "10.1083/jcb.126.3.801",
      "volume": "126",
      "author": "Jalink K.",
      "year": "1994",
      "journal-title": "J. Cell Biol."
    },
    {
      "key": "B17",
      "doi-asserted-by": "crossref",
      "first-page": "2489",
      "DOI": "10.1074/jbc.273.5.2489",
      "volume": "273",
      "author": "Katoh H.",
      "year": "1998",
      "journal-title": "J. Biol. Chem."
    },
    {
      "key": "B18",
      "doi-asserted-by": "crossref",
      "first-page": "28700",
      "DOI": "10.1074/jbc.273.44.28700",
      "volume": "273",
      "author": "Katoh H.",
      "year": "1998",
      "journal-title": "J. Biol. Chem."
    },
    {
      "key": "B19",
      "doi-asserted-by": "crossref",
      "first-page": "1201",
      "DOI": "10.1128/MCB.17.3.1201",
      "volume": "17",
      "author": "Kozma R.",
      "year": "1997",
      "journal-title": "Mol. Cell. Biol."
    },
    {
      "key": "B20",
      "doi-asserted-by": "crossref",
      "first-page": "2417",
      "DOI": "10.1242/jcs.110.19.2417",
      "volume": "110",
      "author": "Kranenburg O.",
      "year": "1997",
      "journal-title": "J. Cell Sci."
    },
    {
      "key": "B21",
      "doi-asserted-by": "crossref",
      "first-page": "519",
      "DOI": "10.1016/S0092-8674(00)81371-9",
      "volume": "87",
      "author": "Lamarche N.",
      "year": "1996",
      "journal-title": "Cell"
    },
    {
      "key": "B22",
      "doi-asserted-by": "crossref",
      "first-page": "5313",
      "DOI": "10.1128/MCB.16.10.5313",
      "volume": "16",
      "author": "Leung T.",
      "year": "1996",
      "journal-title": "Mol. Cell. Biol."
    },
    {
      "key": "B23",
      "doi-asserted-by": "crossref",
      "first-page": "29051",
      "DOI": "10.1074/jbc.270.49.29051",
      "volume": "270",
      "author": "Leung T.",
      "year": "1995",
      "journal-title": "J. Biol. Chem."
    },
    {
      "key": "B24",
      "doi-asserted-by": "crossref",
      "first-page": "183",
      "DOI": "10.1016/S1097-2765(00)80019-2",
      "volume": "1",
      "author": "Manser E.",
      "year": "1998",
      "journal-title": "Mol. Cell"
    },
    {
      "key": "B25",
      "doi-asserted-by": "crossref",
      "first-page": "5638",
      "DOI": "10.1093/emboj/17.19.5638",
      "volume": "17",
      "author": "Mao J.",
      "year": "1998",
      "journal-title": "EMBO J."
    },
    {
      "key": "B26",
      "doi-asserted-by": "crossref",
      "first-page": "2208",
      "DOI": "10.1002/j.1460-2075.1996.tb00574.x",
      "volume": "15",
      "author": "Matsui T.",
      "year": "1996",
      "journal-title": "EMBO J."
    },
    {
      "key": "B27",
      "doi-asserted-by": "crossref",
      "first-page": "168",
      "DOI": "10.1016/S0955-0674(97)80059-2",
      "volume": "9",
      "author": "Moolenaar W.H.",
      "year": "1997",
      "journal-title": "Curr. Opin. Cell Biol."
    },
    {
      "key": "B28",
      "doi-asserted-by": "crossref",
      "first-page": "225",
      "DOI": "10.1242/jcs.108.1.225",
      "volume": "108",
      "author": "Nobes C.D.",
      "year": "1995",
      "journal-title": "J. Cell Sci."
    },
    {
      "key": "B29",
      "doi-asserted-by": "crossref",
      "first-page": "389",
      "DOI": "10.1016/0092-8674(92)90163-7",
      "volume": "70",
      "author": "Ridley A.J.",
      "year": "1992",
      "journal-title": "Cell"
    },
    {
      "key": "B30",
      "doi-asserted-by": "crossref",
      "first-page": "1621",
      "DOI": "10.1016/S0960-9822(02)70785-9",
      "volume": "6",
      "author": "Taylor S.J.",
      "year": "1996",
      "journal-title": "Curr. Biol."
    },
    {
      "key": "B31",
      "doi-asserted-by": "crossref",
      "first-page": "2295",
      "DOI": "10.1101/gad.11.18.2295",
      "volume": "11",
      "author": "Van Aelst L.",
      "year": "1997",
      "journal-title": "Genes & Dev."
    },
    {
      "key": "B32",
      "doi-asserted-by": "crossref",
      "first-page": "797",
      "DOI": "10.1083/jcb.139.3.797",
      "volume": "139",
      "author": "van Leeuwen F.N.",
      "year": "1997",
      "journal-title": "J. Cell Biol."
    },
    {
      "key": "B33",
      "doi-asserted-by": "crossref",
      "first-page": "6741",
      "DOI": "10.1073/pnas.90.14.6741",
      "volume": "90",
      "author": "Xu N.",
      "year": "1993",
      "journal-title": "Proc. Natl. Acad. Sci. USA"
    }
  ],
  "container-title": "Molecular Biology of the Cell",
  "original-title": [],
  "language": "en",
  "deposited": {
    "date-parts": [
      [
        2022,
        2,
        27
      ]
    ],
    "date-time": "2022-02-27T02:39:07Z",
    "timestamp": 1645929547000
  },
  "score": 1,
  "resource": {
    "primary": {
      "URL": "https://www.molbiolcell.org/doi/10.1091/mbc.10.6.1851"
    }
  },
  "subtitle": [],
  "editor": [
    {
      "given": "Mary C.",
      "family": "Beckerle",
      "sequence": "additional",
      "affiliation": []
    }
  ],
  "short-title": [],
  "issued": {
    "date-parts": [
      [
        1999,
        6
      ]
    ]
  },
  "references-count": 33,
  "journal-issue": {
    "issue": "6",
    "published-print": {
      "date-parts": [
        [
          1999,
          6
        ]
      ]
    }
  },
  "alternative-id": [
    "10.1091/mbc.10.6.1851"
  ],
  "URL": "http://dx.doi.org/10.1091/mbc.10.6.1851",
  "relation": {},
  "ISSN": [
    "1059-1524",
    "1939-4586"
  ],
  "subject": [],
  "container-title-short": "MBoC",
  "published": {
    "date-parts": [
      [
        1999,
        6
      ]
    ]
  }
}