INVESTIGADORES
WEINER Andrea Maria Julia
congresos y reuniones científicas
Título:
A Zebrafish model for Richieri-Costa-Pereira Syndrome: knockdown of eif4a3 gene results in craniofacial development disorder
Autor/es:
LUCAS ALVIZI; ANDREA M.J. WEINER; FRANCINE P FAVARO; R. ZECHI-CEIDE; ANTONIO RICHIERI-COSTA; M GUION-ALMEIDA; NORA B CALCATERRA; MARIA RITA PASSOS-BUENO
Lugar:
San Francisco - CA
Reunión:
Congreso; American Society of Human Genetics Meeting; 2012
Resumen:
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Purpose: Richieri-Costa-Pereira Syndrome (RCPS) is a rare acrofacial dysostosis
characterized by Robin sequence, cleft mandible, short stature and limb
defects. RCPS is inherited in an autosomic recesive fashion, presenting a high
rate of consanguinity, increased male mortality and 25% recurrence in sibs
among patient?s families. Recently, our group has identified a mutation in
EIF4A3 gene (Eukariotic Initiation Factor 4A3) in homozygosity in RCPS patients
as a putative molecular cause underlying this disorder (Favaro et al., ASHG
2012 submitted). Once zebrafish
modeling for human genetic diseases has been emerging as a very informative tool
in the developmental field, our objective was to knockdown eif4a3 expression in zebrafish embryos in order to phenocopy RCPS and
study the malformation of craniofacial structures during embryonic development.
Methods: We injected zebrafish
embryos at the two-cells stage with either an eif4a3 translation-blocking or a mismatch control morpholinos (GeneTools).
Development was observed at the points of 4, 15, 24 and 48 hours
post-fertilization (hpf) and 5 days post-fertilization (dpf). Expression of p53 and p21 were assessed through qRT-PCR using 24 hpf embryos to check morpholino
off-target effects and p53 apoptotic
pathway activation by p21. For a deeper
phenotype characterization, whole-mount RNA in situ hybridization for myoD, sox9b and col2a1 expression were performed. Besides, cartilage and
bone formation were analysed by alcian blue and calcein stainings. Finally, by
acridine orange staining we detected the apoptosis level assessment. Results/Conclusion: At 24 hpf, 71% of
embryos in average treated with eif4a3
translation-blocking morpholino presented severely affected as well as
dysmorphic craniofacial structures and a mortality rate of 29%. Forty-eight hpf
embryos were smaller in size than controls and also presented craniofacial
abnormalities. Larvae at 5 dpf had no mandible development. p53 mediated apoptosis pathway was not
overexpressed with morpholino treatment since p53 and p21 expression
was not significantly distinct from negative controls, excluding an off-target
effect from morpholino. Our results suggest that eif4a3 embryonic expression is essential for normal craniofacial
development and its impairment leads to craniofacial dysmorphisms in zebrafish.
Besides, eif4a3 knockdown in zebrafish appears to be a good model for future
research in Richieri-Costa-Pereira Syndrome. CEPID/FAPESP/CONICET
