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RhoA is dispensable for skin development, but crucial for contraction and directed migration of keratinocytes.

Jackson B, Peyrollier K, Pedersen E, Basse A, Karlsson R, Wang Z, Lefever T, Ochsenbein AM, Schmidt G, Aktories K, Stanley A, Quondamatteo F, Ladwein M, Rottner K, van Hengel J, Brakebusch C - Mol. Biol. Cell (2011)

Bottom Line: Rho-inhibiting toxins further increased multinucleation of RhoA- cells but had no significant effect on spreading, suggesting that RhoB and RhoC have partially overlapping functions with RhoA.Loss of RhoA decreased directed cell migration in vitro caused by reduced migration speed and directional persistence.These defects were not related to the decreased cell contraction and were independent of ROCK, as ROCK inhibition by Y27632 increased directed migration of both control and RhoA- keratinocytes.

View Article: PubMed Central - PubMed

Affiliation: Biomedical Institute, BRIC, University of Copenhagen, 2200 Copenhagen, Denmark.

ABSTRACT
RhoA is a small guanosine-5'-triphosphatase (GTPase) suggested to be essential for cytokinesis, stress fiber formation, and epithelial cell-cell contacts. In skin, loss of RhoA was suggested to underlie pemphigus skin blistering. To analyze RhoA function in vivo, we generated mice with a keratinocyte-restricted deletion of the RhoA gene. Despite a severe reduction of cofilin and myosin light chain (MLC) phosphorylation, these mice showed normal skin development. Primary RhoA- keratinocytes, however, displayed an increased percentage of multinucleated cells, defective maturation of cell-cell contacts. Furthermore we observed increased cell spreading due to impaired RhoA-ROCK (Rho-associated protein kinase)-MLC phosphatase-MLC-mediated cell contraction, independent of Rac1. Rho-inhibiting toxins further increased multinucleation of RhoA- cells but had no significant effect on spreading, suggesting that RhoB and RhoC have partially overlapping functions with RhoA. Loss of RhoA decreased directed cell migration in vitro caused by reduced migration speed and directional persistence. These defects were not related to the decreased cell contraction and were independent of ROCK, as ROCK inhibition by Y27632 increased directed migration of both control and RhoA- keratinocytes. Our data indicate a crucial role for RhoA and contraction in regulating cell spreading and a contraction-independent function of RhoA in keratinocyte migration. In addition, our data show that RhoA is dispensable for skin development.

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Stress fibers and focal adhesion form in the absence of RhoA. RhoA- (ko) and control (con) keratinocytes were seeded on glass and stained for filamentous actin (a, a′, c, c′) and paxillin (b, b′, c, c′).
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Figure 7: Stress fibers and focal adhesion form in the absence of RhoA. RhoA- (ko) and control (con) keratinocytes were seeded on glass and stained for filamentous actin (a, a′, c, c′) and paxillin (b, b′, c, c′).

Mentions: To study whether stress fibers are able to form in the absence of RhoA, we isolated keratinocytes from control and RhoA ko mice and cultured them on glass at a low density to avoid cell–cell contact. Cells were fixed 2 d after plating and stained for actin filaments and the focal adhesion protein paxillin. Surprisingly, thick actin bundles anchored in paxillin containing focal adhesions at the cell membrane were found both in control and RhoA- keratinocytes, indicating that RhoA is not essential for the formation of actin stress fibers and focal adhesions (Figure 7), consistent with the finding that their assembly is not only inducible by constitutively active RhoA, but also by RhoB and RhoC (Aspenström et al., 2004).


RhoA is dispensable for skin development, but crucial for contraction and directed migration of keratinocytes.

Jackson B, Peyrollier K, Pedersen E, Basse A, Karlsson R, Wang Z, Lefever T, Ochsenbein AM, Schmidt G, Aktories K, Stanley A, Quondamatteo F, Ladwein M, Rottner K, van Hengel J, Brakebusch C - Mol. Biol. Cell (2011)

Stress fibers and focal adhesion form in the absence of RhoA. RhoA- (ko) and control (con) keratinocytes were seeded on glass and stained for filamentous actin (a, a′, c, c′) and paxillin (b, b′, c, c′).
© Copyright Policy - creative-commons
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3046057&req=5

Figure 7: Stress fibers and focal adhesion form in the absence of RhoA. RhoA- (ko) and control (con) keratinocytes were seeded on glass and stained for filamentous actin (a, a′, c, c′) and paxillin (b, b′, c, c′).
Mentions: To study whether stress fibers are able to form in the absence of RhoA, we isolated keratinocytes from control and RhoA ko mice and cultured them on glass at a low density to avoid cell–cell contact. Cells were fixed 2 d after plating and stained for actin filaments and the focal adhesion protein paxillin. Surprisingly, thick actin bundles anchored in paxillin containing focal adhesions at the cell membrane were found both in control and RhoA- keratinocytes, indicating that RhoA is not essential for the formation of actin stress fibers and focal adhesions (Figure 7), consistent with the finding that their assembly is not only inducible by constitutively active RhoA, but also by RhoB and RhoC (Aspenström et al., 2004).

Bottom Line: Rho-inhibiting toxins further increased multinucleation of RhoA- cells but had no significant effect on spreading, suggesting that RhoB and RhoC have partially overlapping functions with RhoA.Loss of RhoA decreased directed cell migration in vitro caused by reduced migration speed and directional persistence.These defects were not related to the decreased cell contraction and were independent of ROCK, as ROCK inhibition by Y27632 increased directed migration of both control and RhoA- keratinocytes.

View Article: PubMed Central - PubMed

Affiliation: Biomedical Institute, BRIC, University of Copenhagen, 2200 Copenhagen, Denmark.

ABSTRACT
RhoA is a small guanosine-5'-triphosphatase (GTPase) suggested to be essential for cytokinesis, stress fiber formation, and epithelial cell-cell contacts. In skin, loss of RhoA was suggested to underlie pemphigus skin blistering. To analyze RhoA function in vivo, we generated mice with a keratinocyte-restricted deletion of the RhoA gene. Despite a severe reduction of cofilin and myosin light chain (MLC) phosphorylation, these mice showed normal skin development. Primary RhoA- keratinocytes, however, displayed an increased percentage of multinucleated cells, defective maturation of cell-cell contacts. Furthermore we observed increased cell spreading due to impaired RhoA-ROCK (Rho-associated protein kinase)-MLC phosphatase-MLC-mediated cell contraction, independent of Rac1. Rho-inhibiting toxins further increased multinucleation of RhoA- cells but had no significant effect on spreading, suggesting that RhoB and RhoC have partially overlapping functions with RhoA. Loss of RhoA decreased directed cell migration in vitro caused by reduced migration speed and directional persistence. These defects were not related to the decreased cell contraction and were independent of ROCK, as ROCK inhibition by Y27632 increased directed migration of both control and RhoA- keratinocytes. Our data indicate a crucial role for RhoA and contraction in regulating cell spreading and a contraction-independent function of RhoA in keratinocyte migration. In addition, our data show that RhoA is dispensable for skin development.

Show MeSH
Related in: MedlinePlus