Abstract
Polo-like kinase 3 (Plk3) plays an important role in the regulation of cell cycle progression and stress responses. Plk3 also has a tumor-suppressing activity as aging PLK3-null mice develop tumors in multiple organs. The growth of highly vascularized tumors in PLK3-null mice suggests a role for Plk3 in angiogenesis and cellular responses to hypoxia. By studying primary isogenic murine embryonic fibroblasts, we tested the hypothesis that Plk3 functions as a component in the hypoxia signaling pathway. PLK3(-/-) murine embryonic fibroblasts contained an enhanced level of HIF-1α under hypoxic conditions. Immunoprecipitation and pulldown analyses revealed that Plk3 physically interacted with HIF-1α under hypoxia. Purified recombinant Plk3, but not a kinase-defective mutant, phosphorylated HIF-1α in vitro, resulting in a major mobility shift. Mass spectrometry identified two unique serine residues that were phosphorylated by Plk3. Moreover, ectopic expression followed by cycloheximide or pulse-chase treatment demonstrated that phospho-mutants exhibited a much longer half-life than the wild-type counterpart, strongly suggesting that Plk3 directly regulates HIF-1α stability in vivo. Combined, our study identifies Plk3 as a new and essential player in the regulation of the hypoxia signaling pathway.
Highlights
The Polo-like kinase (Plk)3 family is composed of evolutionarily conserved proteins that play an important role in the regulation of cell survival and proliferation, cell cycle progression, and stress responses [1, 2]
To examine the direct regulatory relationship between Polo-like kinase 3 (Plk3) and Hypoxia-inducible factor 1 (HIF-1)␣, we examined their physical interaction during normoxic and hypoxic conditions
Co-immunoprecipitation experiments revealed that the Plk3 antibody, but not a control IgG, pulls down HIF-1␣ in cells treated with hypoxia for 4 h (Fig. 1B)
Summary
The Polo-like kinase (Plk)3 family is composed of evolutionarily conserved proteins that play an important role in the regulation of cell survival and proliferation, cell cycle progression, and stress responses [1, 2]. Polo-like kinase 3 (Plk3) plays an important role in the regulation of cell cycle progression and stress responses. The resin was extensively washed, and proteins that were bound to either His6-Plk3 or the control resin were eluted in the SDS sample buffer and subjected to SDS-PAGE analysis followed by Western blotting with HIF-1␣ antibody.
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
