G-QUADRUPLEXES CONTROL THE EXPRESSION OF DEVELOPMENTALLY REGULATED GENES IN VIVO

DAVID, A.P.; MARGARIT, E.; BANCHIO, C.; CALCATERRA, N.B.; ARMAS, P.

Mar del Plata

Congreso; 51 Annual Meeting Argentine Society for Biochemistry and Molecular Biology; 2015

Argentine Society for Biochemistry and Molecular Biology

Genomic DNA may transiently fold as G-quadruplex (G4), a non-canonical structure associated with gene expression control and genome integrity. Although G4 formation and function was demonstrated in vitro and in cellulo, in vivo biological relevance of this structure is still elusive. To approach this, we used zebrafish embryonic development as an in vivo model to assess the role of G4 on the transcription of conserved developmentally regulated genes. In previous studies we found three developmentally regulated genes containing conserved putative quadruplex sequences (PQS) within their promoters (-1000 bp) that formed G4 in vitro. Here we present in cellulo data obtained by cloning human and zebrafish PQS upstream a basal promoter controlling firefly luciferase transcription in neuro2A cell line, which reveal a transcriptional enhancer role for all tested G4. Furthermore, the in vivo role of these G4 was analyzed by specific disruption microinjecting antisense oligonucleotides in zebrafish embryos. This strategy led to transcriptional reduction of the analyzed genes measured by RT-qPCR and morphological changes previously reported for the loss of function of these genes. Overall, this study indicates that G4 display a function in vivo and may act as conserved gene expression fine-tuning elements during embryonic development.