RESUMEN
The Hippo signaling pathway is a major regulator of organ growth, which controls the activity of the transcription coactivator Yorkie (Yki) in Drosophila and its homolog YAP in mammals. Both Yki and YAP proteins exist as alternatively spliced isoforms containing either one or two WW domains. The biological importance of this conserved alternative splicing event is unknown. Here, we identify the splicing factor B52 as a regulator of yki alternative splicing in Drosophila and show that B52 modulates growth in part through modulation of yki alternative splicing. Yki isoforms differ by their transcriptional activity as well as their ability to bind and bridge PPxY motifs-containing partners, and can compete in vivo. Strikingly, flies in which yki alternative splicing has been abrogated, thus expressing only Yki2 isoform, exhibit fluctuating wing asymmetry, a signal of developmental instability. Our results identify yki alternative splicing as a new level of modulation of the Hippo pathway, that is required for growth equilibration during development. This study provides the first demonstration that the process of alternative splicing contributes to developmental robustness.
Asunto(s)
Proteínas de Drosophila/genética , Proteínas de Drosophila/metabolismo , Drosophila melanogaster/crecimiento & desarrollo , Proteínas Nucleares/genética , Proteínas Nucleares/metabolismo , Factores de Empalme de ARN/metabolismo , Transactivadores/genética , Transactivadores/metabolismo , Empalme Alternativo , Animales , Línea Celular , Proteínas de Drosophila/química , Drosophila melanogaster/genética , Drosophila melanogaster/metabolismo , Regulación del Desarrollo de la Expresión Génica , Técnicas de Silenciamiento del Gen , Proteínas Nucleares/química , Unión Proteica , Dominios Proteicos , Factores de Empalme de ARN/genética , Análisis de Secuencia de ARN , Transactivadores/química , Alas de Animales/crecimiento & desarrollo , Alas de Animales/metabolismo , Proteínas Señalizadoras YAPRESUMEN
Cyclin M1 (CNNM1) functions as a copper storage protein in neuronal cells. We report that Cnnm1 is expressed in mouse testis and brain and has a coding sequence of 1761 bp that encodes a 586 amino acid protein with a molecular weight of 66 kDa. Cnnm1 is expressed in the testes of mice from neonatal to adult stages with relatively higher levels in neonates. CNNM1 expression appeared to be restricted to c-KIT- and OCT3/4-positive cells in the testis, indicating that they are early spermatogonial cells. Spermatogonial stem cells in primary culture expressed Cnnm1, and their differentiation into embryoid body-like clusters in vitro resulted in the loss of Cnnm1 expression. Silencing of Cnnm1 in GC1-spg cells resulted in a significant reduction in the number of cells in G1 phase with concomitant increase in the numbers of cells in both S and G2/M phases. Further, retinoic acid downregulated the expression of Cnnm1 in GC1-spg cells. We conclude that CNNM1 is associated with stemness and self-renewal, and its downregulation triggers differentiation in spermatogonial cells in mouse.