Sipl1 and Rbck1 Are Novel Eya1-Binding Proteins with a Role in Craniofacial Development

Leibniz Institute for Age Research-Fritz Lipmann Institute, Jena, Germany.
Molecular and Cellular Biology (Impact Factor: 4.78). 10/2010; 30(24):5764-75. DOI: 10.1128/MCB.01645-09
Source: PubMed


The eyes absent 1 protein (Eya1) plays an essential role in the development of various organs in both invertebrates and vertebrates. Mutations in the human EYA1 gene are linked to BOR (branchio-oto-renal) syndrome, characterized by kidney defects, hearing loss, and branchial arch anomalies. For a better understanding of Eya1's function, we have set out to identify new Eya1-interacting proteins. Here we report the identification of the related proteins Sipl1 (Shank-interacting protein-like 1) and Rbck1 (RBCC protein interacting with PKC1) as novel interaction partners of Eya1. We confirmed the interactions by glutathione S-transferase (GST) pulldown analysis and coimmunoprecipitation. A first mechanistic insight is provided by the demonstration that Sipl1 and Rbck1 enhance the function of Eya proteins to act as coactivators for the Six transcription factors. Using reverse transcriptase PCR (RT-PCR) and in situ hybridization, we show that Sipl1 and Rbck1 are coexpressed with Eya1 in several organs during embryogenesis of both the mouse and zebrafish. By morpholino-mediated knockdown, we demonstrate that the Sipl1 and Rbck1 orthologs are involved in different aspects of zebrafish development. In particular, knockdown of one Sipl1 ortholog as well as one Rbck1 ortholog led to a BOR syndrome-like phenotype, with characteristic defects in ear and branchial arch formation.

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    • "The reasons for the presumed lack of interaction between Eya1 and Six1a in the neuronal lineage are unknown. However, three possible reasons are: (1) a co-factor for which Eya1 has more affinity than Six1a is present in the neuronal lineage, preventing Eya1 binding to Six1a and allowing Gro1 to bind Six1a in this lineage; (2) either Six1a, Eya1, or both, are modified at the post-translational level (for example, by phosphorylation or methylation) in the neuronal lineage and such modifications prevent their mutual interactions, leaving Six1a available to interact with Gro1; or (3) the Eya1-Six1a complex is more labile in vivo than it is in vitro and such a complex needs to be stabilized by a third partner only expressed in the sensory lineage but absent in the neuronal lineage, such Eya1 co-factors have been recently characterized in mice and zebrafish (Landgraf et al. 2010). "
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