February 7 2007
12:00 MBI 159

Abstract:

The Notch signaling pathway defines an evolutionarily-conserved signal transduction cascade, which plays an essential role in pattern formation and cell-fate determination through local cell-cell interactions. Data from my lab and others have shown that the alternation of cell fate via lateral inhibition is not specific to neurogenesis. Other tissues with several cell types, e.g. sensory patches in the inner ear and pronephros, also use similar mechanisms to specify cell fate. Notch signaling has been also found essential for the ticking of the segmentation clock, which control the periodic generation of somites through oscillating gene expression. Moreover, it is important to demarcate hindbrain boundaries. I will present our recent progress and focus on two stories about a pleiotropic zebrafish mutant, mind bomb (mib), which harbors a mutation in a gene that encodes a RING E3 ligase. Mib has been shown to promote Delta ubiquitylation and endocytosis and, therefore, activate Notch signaling.



We further clone zebrafish mib homologue, mind bomb-2 (mib2). Mib and Mib2 can form homo- and hetero-oligomers and ubiquitylate themselves and each other. Both function similarly in Notch signaling. However, Mib2 is different from Mib in the substrate specificity. DeltaC is a common substrate of Mib and Mib2, while only Mib can promote the internalization of DeltaD. When phenotypes of three available zebrafish mib alleles: mibta52b (missense mutation in the C-terminal RF, M1013R), mibm132 (nonsense mutation results in a truncated protein that loses all the three RFs, C785stop) and mibtfi91 (nonsense mutation that encodes only 60aa protein, Y60stop) are compared, mibta52b mutants have the most severe phenotypes, followed by mibm132 and mibtfi91 mutants. Morpholino (MO) and mRNA injection data indicate that Mibta52b and Mibm132 are dominant-negative forms with a dosage-dependent effect on Mib2, which is the molecular basis for the different phenotypic severity among the mib alleles. Furthermore, we show that Notch activity negatively regulates mib transcription in a Su(h)-dependent manner. mib expression is up-regulated in mibta52b, but not in mibm132 and mibtfi91 embryos.



Two types of epithelial cells, multi-cilia cells and principal cells, were found in the epithelia of zebrafish distal pronephric duct. While the former are characterized with at least 15 apically-localized cilia and expressed centrin2 and rfx2, the latter are characterized with single primary cilia and sodium pumps. Multi-cilia cells and principal cells differentiate between 17.5 to 22 hours post-fertilization in a mosaic pattern. Jagged2a-Notch1a/Notch5-Her9 is responsible for differentiation of these two cell types through a lateral inhibition mechanism. Multi-cilia cell hyperplasia was observed in the mind bomb (mib) mutants. Moreover, Mib interacts with Jagged2a and facilitates its internalization. Taken together, our findings add a new paradigm of Notch signaling in kidney development, namely, cell fate differentiation in a nephric segment (distal pronephric duct) is modulated by Jagged2a-Notch signaling.