Purpose: : Progressive specification of the eye structures, during vertebrate embryonic development, requires precise regulation of morphogenetic events, which are coordinated by networks of transcription factors and cell signalling pathways. Regulation of gene activity by microRNAs (miRNAs) is critical in multiple aspects of vertebrate embryonic development. Here, we focus on the in vivo role of miR-204, which is highly expressed in the developing retinal pigment epithelium, lens and retina. Methods: : miR-204 function, during Oryzias latipes (medaka fish) eye development, was assessed by injection of antisense "multiblocking" morpholino oligonucleotides, which specifically prevented biogenesis and activity of this microRNA. Heteroduplex miR-204 oligonucleotides injections were performed to increase miR-204 activity. Analysis of eye phenotypes in injected embryos included morphological inspection and molecular analyses by RNA in situ hybridization and immunohistochemistry. Results: : The knockdown of miR-204 in medaka fish led to a specific eye phenotype, characterised by microphthalmia, coloboma and aberrant lens formation. In addition, disruption of miR-204 activity determined significant defects in ventral retina cell differentiation processes. We demonstrated that the miR-204-morphant eye phenotype is in part due to increased cell death rather than decreased proliferation. We showed that miR-204 acts by directly repressing positive regulators of the PAX6 gene. Finally, miR-204 over-expression led to changes in the expression levels of retinal and lens cell specific markers, which were mostly complementary to those observed following miR-204 inactivation. Conclusions: : Taken together, these results not only demonstrate that a single miRNA can regulate the folding of the optic vesicle to form the optic cup and the formation of a mature and functional eye, but also establish the existence of a precise network between miR-204 and Pax6 in mediating retina and lens development.
Mir-204 Modulates Optic Cup Patterning During Medaka Fish Embryonic Eye Development
Karali M;
2009
Abstract
Purpose: : Progressive specification of the eye structures, during vertebrate embryonic development, requires precise regulation of morphogenetic events, which are coordinated by networks of transcription factors and cell signalling pathways. Regulation of gene activity by microRNAs (miRNAs) is critical in multiple aspects of vertebrate embryonic development. Here, we focus on the in vivo role of miR-204, which is highly expressed in the developing retinal pigment epithelium, lens and retina. Methods: : miR-204 function, during Oryzias latipes (medaka fish) eye development, was assessed by injection of antisense "multiblocking" morpholino oligonucleotides, which specifically prevented biogenesis and activity of this microRNA. Heteroduplex miR-204 oligonucleotides injections were performed to increase miR-204 activity. Analysis of eye phenotypes in injected embryos included morphological inspection and molecular analyses by RNA in situ hybridization and immunohistochemistry. Results: : The knockdown of miR-204 in medaka fish led to a specific eye phenotype, characterised by microphthalmia, coloboma and aberrant lens formation. In addition, disruption of miR-204 activity determined significant defects in ventral retina cell differentiation processes. We demonstrated that the miR-204-morphant eye phenotype is in part due to increased cell death rather than decreased proliferation. We showed that miR-204 acts by directly repressing positive regulators of the PAX6 gene. Finally, miR-204 over-expression led to changes in the expression levels of retinal and lens cell specific markers, which were mostly complementary to those observed following miR-204 inactivation. Conclusions: : Taken together, these results not only demonstrate that a single miRNA can regulate the folding of the optic vesicle to form the optic cup and the formation of a mature and functional eye, but also establish the existence of a precise network between miR-204 and Pax6 in mediating retina and lens development.I documenti in IRIS sono protetti da copyright e tutti i diritti sono riservati, salvo diversa indicazione.
