RESUMO
Focal adhesion turnover during cell migration is an integrated cyclic process requiring tight regulation of integrin function. Interaction of integrin with its ligand depends on its activation state, which is regulated by the direct recruitment of proteins onto the ß integrin chain cytoplasmic domain. We previously reported that ICAP-1α, a specific cytoplasmic partner of ß1A integrins, limits both talin and kindlin interaction with ß1 integrin, thereby restraining focal adhesion assembly. Here we provide evidence that the calcium and calmodulin-dependent serine/threonine protein kinase type II (CaMKII) is an important regulator of ICAP-1α for controlling focal adhesion dynamics. CaMKII directly phosphorylates ICAP-1α and disrupts an intramolecular interaction between the N- and the C-terminal domains of ICAP-1α, unmasking the PTB domain, thereby permitting ICAP-1α binding onto the ß1 integrin tail. ICAP-1α direct interaction with the ß1 integrin tail and the modulation of ß1 integrin affinity state are required for down-regulating focal adhesion assembly. Our results point to a molecular mechanism for the phosphorylation-dependent control of ICAP-1α function by CaMKII, allowing the dynamic control of ß1 integrin activation and cell adhesion.
Assuntos
Proteína Quinase Tipo 2 Dependente de Cálcio-Calmodulina/metabolismo , Adesões Focais/metabolismo , Integrina beta1/metabolismo , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Animais , Benzilaminas/farmacologia , Células CHO , Proteína Quinase Tipo 2 Dependente de Cálcio-Calmodulina/antagonistas & inibidores , Proteína Quinase Tipo 2 Dependente de Cálcio-Calmodulina/genética , Adesão Celular/efeitos dos fármacos , Movimento Celular/efeitos dos fármacos , Células Cultivadas , Cricetinae , Cricetulus , Adesões Focais/efeitos dos fármacos , Adesões Focais/genética , Immunoblotting , Integrina beta1/genética , Peptídeos e Proteínas de Sinalização Intracelular/genética , Camundongos , Camundongos Knockout , Microscopia Confocal , Modelos Biológicos , Mutação , Células NIH 3T3 , Osteoblastos/citologia , Osteoblastos/efeitos dos fármacos , Osteoblastos/metabolismo , Fosforilação , Inibidores de Proteínas Quinases/farmacologia , Ratos , Sulfonamidas/farmacologia , Treonina/genética , Treonina/metabolismo , Imagem com Lapso de TempoRESUMO
BACKGROUND: The corneal epithelium (CE) overlays a stroma, which is derived from neural crest cells, and appears to be committed during chick development, but appears still labile in adult rabbit. Its specification was hitherto regarded as resolved and dependent upon the lens, although without experimental support. Here, we challenged CE fate by changing its environment at different stages. RESULTS: Recombination with a dermis showed that CE commitment is linked to stroma formation, which results in Pax6 stabilization in both species. Surgical ablation shows that CE specification has already taken place when the lens placode invaginates, while removal of the early lens placode led to lens renewal. To block lens formation, bone morphogenetic protein (BMP) signaling, one of its last inducing factors, was inhibited by over-expression of Gremlin in the ocular ectoderm. This resulted in lens-less embryos which formed a corneal epithelium if they survived 2 weeks. CONCLUSION: The corneal epithelium and lens share a common pool of precursors. The adoption of the CE fate might be dependent on the loss of a lens placode favoring environment. The corneal fate is definitively stabilized by the migration of Gremlin-expressing neural crest cells in the lens peripheral ectoderm.
Assuntos
Epitélio Corneano/embriologia , Cristalino/embriologia , Células-Tronco/fisiologia , Animais , Animais Geneticamente Modificados , Diferenciação Celular/genética , Diferenciação Celular/fisiologia , Linhagem da Célula/genética , Linhagem da Célula/fisiologia , Movimento Celular/genética , Movimento Celular/fisiologia , Embrião de Galinha , Ectoderma/citologia , Ectoderma/embriologia , Ectoderma/metabolismo , Ectoderma/fisiologia , Epitélio Corneano/citologia , Epitélio Corneano/crescimento & desenvolvimento , Epitélio Corneano/metabolismo , Proteínas do Olho/genética , Proteínas do Olho/metabolismo , Regulação da Expressão Gênica no Desenvolvimento , Proteínas de Homeodomínio/genética , Proteínas de Homeodomínio/metabolismo , Cristalino/citologia , Cristalino/crescimento & desenvolvimento , Cristalino/metabolismo , Células-Tronco Mesenquimais/metabolismo , Células-Tronco Mesenquimais/fisiologia , Modelos Biológicos , Fator de Transcrição PAX6 , Fatores de Transcrição Box Pareados/genética , Fatores de Transcrição Box Pareados/metabolismo , Coelhos , Proteínas Repressoras/genética , Proteínas Repressoras/metabolismo , Células-Tronco/metabolismoRESUMO
Feathers, like hairs, first appear as primordia consisting of an epidermal placode associated with a dermal condensation that is necessary for the continuation of their differentiation. Previously, the BMPs have been proposed to inhibit skin appendage formation. We show that the function of specific BMPs during feather development is more complex. BMP2 and BMP7, which are expressed in both the epidermis and the dermis, are involved in an antagonistic fashion in regulating the formation of dermal condensations, and thus are both necessary for subsequent feather morphogenesis. BMP7 is expressed earlier and functions as a chemoattractant that recruits cells into the condensation, whereas BMP2 is expressed later, and leads to an arrest of cell migration, likely via its modulation of the EIIIA fibronectin domain and alpha4 integrin expression. Based on the observed cell proliferation, chemotaxis and the timing of BMP2 and BMP7 expression, we propose a mathematical model, a reaction-diffusion system, which not only simulates feather patterning, but which also can account for the negative effects of excess BMP2 or BMP7 on feather formation.
