RESUMO
Recent experiments suggest that Xenopus Neurotrophin Receptor Homolog 1 (NRH1) proteins act through the planar cell polarity pathway to regulate convergent extension movements during gastrulation and neurulation. We show in this paper that NRH1 proteins are also required for the proper expression of mesodermally expressed genes such as Xbra and Chordin, and to a lesser extent, of Xwnt11. Loss of NRH1 function is followed, during gastrula and neurula stages, by a dramatic increase in apoptosis. Apoptosis is delayed by injection of Xbra RNA, suggesting that cell death is a consequence, at least in part, of the down-regulation of this gene, and it is also delayed by expression of activated forms of Rho, Rac and Cdc42. These small GTPases have previously been implicated in the planar cell polarity pathway in Xenopus and, in other systems, in the regulation of apoptosis. We conclude that the effects of NRH1 proteins include the regulation of mesodermal gene expression and that the disruption of gastrulation that is caused by their loss of function is a consequence of the down-regulation of Xbra and other genes, in addition to direct interference with the planar cell polarity pathway. The apoptosis observed in embryos lacking NRH1 function is not an indirect consequence of the disruption of gastrulation, and indeed it may contribute to the observed morphological defects.
Assuntos
Apoptose/fisiologia , Embrião não Mamífero/fisiologia , Gástrula/fisiologia , Mesoderma/fisiologia , Receptores de Fator de Crescimento Neural/fisiologia , Proteínas de Xenopus/fisiologia , Xenopus laevis/embriologia , Animais , Sequência de Bases , Glicoproteínas/fisiologia , Peptídeos e Proteínas de Sinalização Intercelular/fisiologia , Dados de Sequência Molecular , Oligonucleotídeos Antissenso , Receptores de Fator de Crescimento Neural/antagonistas & inibidores , Receptores de Fator de Crescimento Neural/biossíntese , Receptores de Fator de Crescimento Neural/genética , Proteínas com Domínio T/fisiologia , Proteínas Wnt/fisiologia , Proteínas de Xenopus/genética , Proteínas rho de Ligação ao GTP/biossíntese , Proteínas rho de Ligação ao GTP/genéticaRESUMO
Members of the T-box gene family play important and diverse roles in development and disease. Here, we study the functional specificities of the Xenopus T-domain proteins Xbra and VegT, which differ in their abilities to induce gene expression in prospective ectodermal tissue. In particular, VegT induces strong expression of goosecoid whereas Xbra cannot. Our results indicate that Xbra is unable to induce goosecoid because it directly activates expression of Xom, a repressor of goosecoid that acts downstream of BMP signaling. We show that the inability of Xbra to induce goosecoid is imposed by an N-terminal domain that interacts with the C-terminal MH2 domain of Smad1, a component of the BMP signal transduction pathway. Interference with this interaction causes ectopic activation of goosecoid and anteriorization of the embryo. These findings suggest a mechanism by which individual T-domain proteins may interact with different partners to elicit a specific response.
Assuntos
Proteínas de Ligação a DNA/metabolismo , Regulação da Expressão Gênica no Desenvolvimento , Morfogênese , Proteínas com Domínio T/metabolismo , Transativadores/metabolismo , Proteínas de Xenopus/metabolismo , Xenopus laevis/fisiologia , Sequência de Aminoácidos , Animais , Proteínas Morfogenéticas Ósseas/metabolismo , Proteínas de Ligação a DNA/química , Proteínas de Ligação a DNA/genética , Ectoderma/citologia , Ectoderma/fisiologia , Proteína Goosecoid , Proteínas de Homeodomínio/genética , Proteínas de Homeodomínio/metabolismo , Humanos , Modelos Moleculares , Dados de Sequência Molecular , Conformação Proteica , Estrutura Terciária de Proteína , Proteínas Repressoras/genética , Proteínas Repressoras/metabolismo , Alinhamento de Sequência , Transdução de Sinais/fisiologia , Proteínas Smad , Proteína Smad1 , Proteínas com Domínio T/química , Proteínas com Domínio T/genética , Transativadores/química , Transativadores/genética , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo , Técnicas do Sistema de Duplo-Híbrido , Proteínas de Xenopus/genética , Xenopus laevis/embriologiaRESUMO
Members of the Bix family of homeobox-containing genes are expressed in the vegetal hemisphere of the Xenopus embryo at the early gastrula stage. Misexpression of at least some of the family members causes activation of mesoderm- and endoderm-specific genes and it is known that some of the proteins, including Bix2 and Bix3, interact with Smad proteins via a motif that is also present in the related protein Mixer. In this paper we study the function of Bix3. Misexpression of Bix3, similar to misexpression of other members of the Bix family, causes the activation of a range of mesendodermal genes, but the spectrum of genes induced by Bix3 differs from that induced by Bix1. More significantly, we find that overexpression of Bix3 also causes apoptosis, as does depletion of Bix3 by use of antisense morpholino oligonucleotides. The ability of Bix3 to causes apoptosis is not associated with its ability to activate transcription and nor with its possession of a Smad interaction motif. Rather, Bix3 lacks a C-terminal motif, which, in Bix1, acts in cis to inhibit apoptosis. Mutation of this sequence in Bix1 causes the protein to acquire apoptosis-inducing activity.
