Journal article
The Functional Significance of Posttranslational Modifications on Polo-Like Kinase 1 Revealed by Chemical Genetic Complementation
PloS one, Vol.11(2), pp.e0150225-e0150225
02/26/2016
DOI: 10.1371/journal.pone.0150225
PMCID: PMC4769148
PMID: 26919439
Abstract
Mitosis is coordinated by carefully controlled phosphorylation and ubiquitin-mediated proteolysis. Polo-like kinase 1 (Plk1) plays a central role in regulating mitosis and cytokinesis by phosphorylating target proteins. Yet, Plk1 is itself a target for posttranslational modification by phosphorylation and ubiquitination. We developed a chemical-genetic complementation assay to evaluate the functional significance of 34 posttranslational modifications (PTMs) on human Plk1. To do this, we used human cells that solely express a modified analog-sensitive Plk1 (Plk1AS) and complemented with wildtype Plk1. The wildtype Plk1 provides cells with a functional Plk1 allele in the presence of 3-MB-PP1, a bulky ATP-analog inhibitor that specifically inhibits Plk1AS. Using this approach, we evaluated the ability of 34 singly non-modifiable Plk1 mutants to complement Plk1AS in the presence of 3-MB-PP1. Mutation of the T-loop activating residue T210 and adjacent T214 are lethal, but surprisingly individual mutation of the remaining 32 posttranslational modification sites did not disrupt the essential functions of Plk1. To evaluate redundancy, we simultaneously mutated all phosphorylation sites in the kinase domain except for T210 and T214 or all sites in the C-terminal polo-box domain (PBD). We discovered that redundant phosphorylation events within the kinase domain are required for accurate chromosome segregation in anaphase but those in the PBD are dispensable. We conclude that PTMs within the T-loop of Plk1 are essential and nonredundant, additional modifications in the kinase domain provide redundant control of Plk1 function, and those in the PBD are dispensable for essential mitotic functions of Plk1. This comprehensive evaluation of Plk1 modifications demonstrates that although phosphorylation and ubiquitination are important for mitotic progression, many individual PTMs detected in human tissue may have redundant, subtle, or dispensable roles in gene function.
Details
- Title: Subtitle
- The Functional Significance of Posttranslational Modifications on Polo-Like Kinase 1 Revealed by Chemical Genetic Complementation
- Creators
- Amber L Lasek - University of Wisconsin–MadisonBrittany M McPherson - University of Wisconsin–MadisonNatalie G Trueman - University of Wisconsin–MadisonMark E Burkard - University of Wisconsin–Madison
- Resource Type
- Journal article
- Publication Details
- PloS one, Vol.11(2), pp.e0150225-e0150225
- DOI
- 10.1371/journal.pone.0150225
- PMID
- 26919439
- PMCID
- PMC4769148
- NLM abbreviation
- PLoS One
- ISSN
- 1932-6203
- eISSN
- 1932-6203
- Grant note
- T32 CA009135 / NCI NIH HHS UL1 TR000427 / NCATS NIH HHS R01 GM097245 / NIGMS NIH HHS 9U54TR000021 / NCATS NIH HHS P30 CA014520 / NCI NIH HHS
- Language
- English
- Date published
- 02/26/2016
- Academic Unit
- Internal Medicine
- Record Identifier
- 9984700652002771
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