Regulation of antagonistic signaling components by miR-430 microRNAs

Cell signaling events that direct animal development are very dose-sensitive processes, and improper regulation leads to abnormal development and disease. My thesis research has found a role for microRNAs (miRNAs) in regulating the dosage of ligands in several signaling pathways. MiRNAs post-transcriptionally repress hundreds of target messenger RNAs (mRNAs), but the physiological roles of specific miRNA-mRNA interactions remain largely elusive. We have focused on one abundantly expressed miRNA family during zebrafish embryonic development. Zebrafish microRNA-430 (miR-430) is activated at the onset of zygotic transcription and directly regulates hundreds of target mRNAs by accelerating deadenylation during the maternal-to-zygotic transition. We have found that miR-430 may also act as a layer of regulation over the Nodal signaling and SDF-1/CXCR4/CXCR7 chemokine signaling pathways during zebrafish development by targeting antagonistic components of each pathway. We have found that the mRNAs encoding the Nodal ligand squint (sqt) and the antagonist lefty2 (lft2) are direct targets of miR-430. Additionally, the chemokine sdf-1a and its decoy receptor cxcr7 are targets of miR-430. To study the effects of each specific targeting event, we developed target protector morpholinos (TPs) that compete with miR-430 for binding to target sites in the 3' UTRs of the mRNAs, allowing us to examine the effect of miR-430 regulation on endogenous targets. Protection of sqt and lft2 mRNAs from miR-430 caused an imbalance and reduction in Nodal signaling. Protection of sdf-1a and cxcr7 mRNAs from miR-430 resulted in aberrant migration of endodermal, trigeminal ganglion, and primordial germ cells. My work suggests that miR-430 regulation of the Nodal signaling and SDF-1/CXCR4/CXCR7 chemokine signaling pathways maintains the correct level of signaling for normal development. To accomplish this, miR-430 fine-tunes the expression of the ligands sqt, lft2, and sdf-1a, which are known to act in concentration-dependent manners. The miR-430-mediated repression of these targets may also act as a buffer against noise in gene expression to increase developmental robustness. Taken together, my thesis work suggest that microRNAs are effective regulators of dosage-sensitive processes during development, and manipulation of miRNA targeting events could lead to potentially therapeutic changes in gene dosage.