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Analysis of the Drosophila Ajuba LIM protein defines functions for distinct LIM domains
Cordelia Rauskolb1, Ahri Han1, Elmira Kirichenko1
1Waksman Institute and Department of Molecular Biology and Biochemistry, Rutgers University, Piscataway, NJ, United States of America.
Insights
The Ajuba LIM protein Jub regulates Hippo signaling and junctional tension. Its distinct LIM domains control interactions with Warts and Steppke, impacting wing growth and cell shape.
Area of Science:
- Cell Biology
- Developmental Biology
- Biochemistry
Background:
- The Ajuba LIM protein, Jub, is crucial for regulating Hippo signaling via cytoskeletal tension and feedback regulation of junctional tension.
- Jub interacts with the kinase Warts and the cytohesin Steppke to mediate these processes.
Purpose of the Study:
- To investigate how Jub interacts with and regulates its distinct partners, Warts and Steppke.
- To determine the specific roles of Jub's three LIM domains in these interactions and regulatory functions.
Main Methods:
- Analyzing the ability of Jub proteins with deleted LIM domains to rescue 'jub' phenotypes in Drosophila wing imaginal discs.
- Performing co-immunoprecipitation experiments in cultured cells and in vivo studies to assess protein interactions and localization.
Main Results:
- Multiple Jub regions bind α-catenin and localize to adherens junctions.
- LIM2 is specifically required for Warts binding in vitro, but LIM1 and LIM2 are needed in vivo for wing growth and Warts localization.
- LIM2 and LIM3 are essential for regulating cell shape and Steppke localization in vivo and for maximal Steppke binding.
Conclusions:
- Distinct LIM domains of the Ajuba LIM protein Jub mediate specific interactions with Warts and Steppke.
- These domain-specific interactions underpin Jub's diverse roles in regulating Hippo signaling, cytoskeletal tension, and developmental processes.
Abstract:
The Ajuba LIM protein Jub mediates regulation of Hippo signaling by cytoskeletal tension through interaction with the kinase Warts and participates in feedback regulation of junctional tension through regulation of the cytohesin Steppke. To investigate how Jub interacts with and regulates its distinct partners, we investigated the ability of Jub proteins missing different combinations of its three LIM domains to rescue jub phenotypes and to interact with α-catenin, Warts and Steppke. Multiple regions of Jub contribute to its ability to bind α-catenin and to localize to adherens junctions in Drosophila wing imaginal discs. Co-immunoprecipitation experiments in cultured cells identified a specific requirement for LIM2 for binding to Warts. However, in vivo, both LIM1 and LIM2, but not LIM3, were required for regulation of wing growth, Yorkie activity, and Warts localization. Conversely, LIM2 and LIM3, but not LIM1, were required for regulation of cell shape and Steppke localization in vivo, and for maximal Steppke binding in co-immunoprecipitation experiments. These observations identify distinct functions for the different LIM domains of Jub.
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