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Roles for alpha 1 connexin in morphogenesis of chick embryos revealed using a novel antisense approach
D L Becker1, I McGonnell, H P Makarenkova
1Department of Anatomy and Developmental Biology, University College London, UK. d.becker@ucl.ac.uk
Insights
Alpha 1 connexin (Cx43) is crucial for embryonic development and pattern formation. Regulating its expression with antisense deoxyoligonucleotides (ODNs) in chick embryos caused severe congenital abnormalities, highlighting Cx43
Area of Science:
- Developmental Biology
- Cellular Biology
- Molecular Biology
Background:
- Gap junctional communication is vital for embryonic development and pattern formation.
- Alpha 1 connexin (Cx43) expression is dynamic and spatially restricted during chick embryogenesis, correlating with developmental events like budding and tissue fusion.
- Cx43 is highly expressed in regions undergoing shaping and tissue fusion.
Purpose of the Study:
- To investigate the role of alpha 1 connexin (Cx43) in embryonic morphogenetic events.
- To develop and utilize a novel method for regulating Cx43 expression in developing chick embryos.
- To understand the impact of Cx43 knockdown on embryonic development and pattern formation.
Main Methods:
- Development of a novel method using unmodified antisense deoxyoligonucleotides (ODNs) delivered via pluronic gel.
- Spatially and temporally regulated knockdown of alpha 1 connexin (Cx43) protein expression in chick embryos.
- Observation and documentation of developmental defects resulting from Cx43 knockdown.
Main Results:
- Cx43 knockdown resulted in significant developmental defects mimicking human congenital abnormalities.
- Observed defects included spina bifida, anencephaly, myeloschisis, limb malformations, cleft palate, and cardiovascular deformities.
- Failure of hematopoiesis was also noted in Cx43-deficient embryos.
Conclusions:
- Alpha 1 connexin (Cx43) plays a major role in integrating signaling pathways essential for embryonic pattern formation.
- Cx43-mediated gap junction communication is critical for normal embryonic development.
- The novel antisense ODN delivery technique using pluronic gel is potentially widely applicable for studying gene function in development.
Abstract:
Gap junctional communication has been implicated in embryonic development and pattern formation. The gap junction protein, alpha 1 connexin (Cx43) is expressed in dynamic and spatially restricted patterns in the developing chick embryo and its expression correlates with many specific developmental events. High levels of expression are found in regions of budding, which leads to shaping and appears to be a necessary prelude for tissue fusions. In order to investigate the role of alpha 1 connexin in these morphogenetic events, we developed a novel method of applying unmodified antisense deoxyoligonucleotides (ODNs) to chick embryos. The use of pluronic gel to deliver antisense ODNs has allowed us to regulate the expression of alpha 1 connexin protein, both spatially and temporally. This "knockdown" results in some striking developmental defects that mimic some common congenital abnormalities, such as spina bifida, anencephaly, myeloschisis, limb malformation, cleft palate, failure of hematopoiesis, and cardiovascular deformity. The results imply a major role for alpha 1 connexin communication in the integration of signaling required for pattern formation during embryonic development. This novel antisense technique may also be widely applicable.