DYNAMIC INTERACTIONS OF NODAL FACTORS AND THEIR RECEPTORS IN ZEBRAFISH EMBRYOS

Abstract

IN MULTICELLULAR ORGANISMS, THE GRADED DISTRIBUTION OF SIGNALING PROTEINS IS REQUIRED FOR MANY DEVELOPMENTAL PROCESSES. THE NODAL SIGNALING PROTEINS, PLAY CRITICAL ROLES IN EMBRYONIC AXIS FORMATION. ZEBRAFISH NODAL FACTORS, SQUINT (SQT) AND CYCLOPS (CYC), HAVE OVERLAPPING FUNCTIONS IN EMBRYOS, BUT DIFFER IN THEIR SIGNALING ACTIVITIES: SQT ACTS AT LONG-RANGE WHEREAS CYC ONLY AFFECTS ADJACENT CELLS. TO EXPLAIN THE DIFFERENCES UNDERLYING SQT AND CYC SIGNALING, THE BIOPHYSICAL PROPERTIES OF SQT AND CYC WERE DETERMINED. THE RESULTS SUGGEST THAT DIFFERENTIAL STABILITY IS A MAJOR CONTRIBUTOR TO NODAL ACTIVITY. MOREOVER, TO BETTER ANALYZE THE FUNCTIONAL DIFFERENCES OF NODALS IN ZEBRAFISH, A METHOD THAT UTILIZES COMBINATIONS OF TARGETED NUCLEASES TO GENERATE LARGE DELETIONS AND RNA NULL MUTANTS WAS DEVELOPED. WITH THIS GENOME EDITING METHOD, CYC AND SQT RNA NULL MUTANTS WERE GENERATED. WHILE GENERATING THESE MUTANTS, SURPRISINGLY, IT WAS FOUND THAT NUCLEASE COMBINATIONS HAVE A SIGNIFICANTLY HIG