Genomic screens: The TAO1 of mitotic checkpoint signaling
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A functional genomic screen using short hairpin RNA (shRNA) libraries directed against kinases and phosphatases has identified a role for the serine/threonine kinase TAO1 in the mitotic spindle checkpoint.
The spindle checkpoint is responsible for delaying the mitotic metaphase-anaphase transition until all chromosomes are correctly aligned at the metaphase plate and attached to microtubule spindles. As such, activation of the checkpoint is critical for avoiding aneuploidy. Several signaling cascades impinge upon the checkpoint and research has been directed towards elucidating the members of these pathways. In Nature Cell Biology, Elledge and colleagues report the use of a short hairpin RNA (shRNA)-based genomic screen to identify novel pathway proteins and uncover a new role for TAO1 (also know as MARKK) in promoting spindle checkpoint activation.
A high-throughput genomic screen with 2500 shRNA sequences was used to target 780 kinase and phosphatase genes. The screen validated known checkpoint kinases such as Bub1, BubR1 and AuroraB, but also identified several proteins with previously unknown roles in checkpoint regulation, among them the serine/threonine kinase TAO1. Although TAO1 has previously been implicated in regulation of microtubule dynamics and p38 MAPK signaling, the authors have now found that the kinase function of TAO1 is critical for spindle checkpoint activation. Coincident with this role, the activity and protein levels of TAO1 were upregulated prior to the anaphase-metaphase transition, and fell as cells completed mitosis.
Small interfering RNA-based depletion of TAO1 resulted in the failure of spindle checkpoint activation. Consequently, anaphase was initiated before chromosomes were properly aligned, causing aberrant chromosomal segregation. TAO1 depletion also caused mislocalization of the checkpoint protein Mad2, which is normally recruited to the site of microtubule attachment at kinetochores. Like TAO1, BubR1 also controls Mad2 localization, and TAO1 and BubR1 were shown to interact in vivo. Therefore, it is likely that TAO1 and BubR1 collaborate to promote Mad2 recruitment to kinetochores and regulate checkpoint signaling. However, it remains unclear whether Mad2 is a direct target for TAO1.
The documented role of TAO1 in regulating microtubule dynamics suggests that it may also govern correct kinetochore attachment through the regulation of microtubule-associated regulatory kinases (MARKs). Aneuploidy is a common trait in cancerous cells, and further study of the biological mechanism behind TAO1-mediated spindle checkpoint activation will aid understanding of the development of genomic instability. In Nature (Nature446, 876-881 (19 April 2007)), Elledge and colleagues described the use of a shRNA-based screen to target ubiquitin-proteasome pathway genes and discovered a role for the deubiquitinating enzyme USP44 in regulating the spindle checkpoint. Clearly, shRNA-based screens hold great promise for uncovering new members of important cell signaling pathways.
Emily J. Chenette Signaling Gateway
References
Viji M. Draviam, Frank Stegmeier, Grzegorz Nalepa, Mathew E. Sowa, Jing Chen, Anthony Liang, Gregory J. Hannon, Peter K. Sorger, J. Wade Harper and Stephen J. Elledge. A functional genomic screen identifies a role for TAO1 kinase in spindle-checkpoint signalling. Nature Cell Biology9, 556-564 (2007) | Article | PubMed |
