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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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Related in: MedlinePlus

RhoA is not crucial for skin wound closure in vivo. In vivo wound closure was analyzed by full thickness wounds in the back skin of control (con) and RhoA fl/fl K5 Cre (ko) mice. Hematoxylin and eosin–staining was used to identify the wound borders in bisectioned wounds (indicated by arrows). Three days after wounding, wound diameters were not significantly different in control and mutant mice (six wounds, three mice). Insets show magnifications of the wound edges. (B) Normal levels of GTP-bound active Rac1and Cdc42 in freshly isolated RhoA- keratinocytes (n = 3/3).
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Figure 10: RhoA is not crucial for skin wound closure in vivo. In vivo wound closure was analyzed by full thickness wounds in the back skin of control (con) and RhoA fl/fl K5 Cre (ko) mice. Hematoxylin and eosin–staining was used to identify the wound borders in bisectioned wounds (indicated by arrows). Three days after wounding, wound diameters were not significantly different in control and mutant mice (six wounds, three mice). Insets show magnifications of the wound edges. (B) Normal levels of GTP-bound active Rac1and Cdc42 in freshly isolated RhoA- keratinocytes (n = 3/3).

Mentions: The impairment of migration of RhoA- keratinocytes in vitro suggested that RhoA function might contribute to skin wound healing in vivo. We therefore tested skin wound healing in RhoA mutant and control mice by making two full-thickness wounds in the back skin of each mouse. Three and five days after wounding mice were sacrificed and the wound gap was measured on hematoxylin and eosin–stained skin sections. In 3 d wounds, no difference was observed between control and mutant mice (Figure 10A). Five days after wounding, wounds were completely closed in both control and RhoA mutant mice (unpublished data).


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)

RhoA is not crucial for skin wound closure in vivo. In vivo wound closure was analyzed by full thickness wounds in the back skin of control (con) and RhoA fl/fl K5 Cre (ko) mice. Hematoxylin and eosin–staining was used to identify the wound borders in bisectioned wounds (indicated by arrows). Three days after wounding, wound diameters were not significantly different in control and mutant mice (six wounds, three mice). Insets show magnifications of the wound edges. (B) Normal levels of GTP-bound active Rac1and Cdc42 in freshly isolated RhoA- keratinocytes (n = 3/3).
© Copyright Policy - creative-commons
Related In: Results  -  Collection

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

Figure 10: RhoA is not crucial for skin wound closure in vivo. In vivo wound closure was analyzed by full thickness wounds in the back skin of control (con) and RhoA fl/fl K5 Cre (ko) mice. Hematoxylin and eosin–staining was used to identify the wound borders in bisectioned wounds (indicated by arrows). Three days after wounding, wound diameters were not significantly different in control and mutant mice (six wounds, three mice). Insets show magnifications of the wound edges. (B) Normal levels of GTP-bound active Rac1and Cdc42 in freshly isolated RhoA- keratinocytes (n = 3/3).
Mentions: The impairment of migration of RhoA- keratinocytes in vitro suggested that RhoA function might contribute to skin wound healing in vivo. We therefore tested skin wound healing in RhoA mutant and control mice by making two full-thickness wounds in the back skin of each mouse. Three and five days after wounding mice were sacrificed and the wound gap was measured on hematoxylin and eosin–stained skin sections. In 3 d wounds, no difference was observed between control and mutant mice (Figure 10A). Five days after wounding, wounds were completely closed in both control and RhoA mutant mice (unpublished data).

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