Supplementary Materials

Supplementary Materials for:

Quantitative Phosphoproteomics Identifies Substrates and Functional Modules of Aurora and Polo-Like Kinase Activities in Mitotic Cells

Arminja N. Kettenbach, Devin K. Schweppe, Brendan K. Faherty, Dov Pechenick, Alexandre A. Pletnev, Scott A. Gerber*

*To whom correspondence should be addressed. E-mail: scott.a.gerber{at}

This PDF file includes:

  • Materials and Methods
  • Fig. S1. Spindle and chromosome morphology of inhibitor-treated HeLa cells.
  • Fig. S2. Plk1 and Plk4 inhibition by BI2536.
  • Fig. S3. Experimental setup and schematic of time line of HeLa cell synchronization.
  • Fig. S4. Flow cytometry analysis of inhibitor-treated HeLa cells.
  • Fig. S5. Ratio distributions.
  • Fig. S6. Comparison of the heavy-to-light ratios in AZD1152- and ZM447439-treated cells.
  • Fig. S7. Candidate Aurora kinase A versus B targets.
  • Fig. S8. Cluster and motif analysis of ambiguous Aurora substrates.
  • Fig. S9. Log2 ratio distribution of known Aurora kinase A, Aurora kinase B, and Plk1 substrates identified in this analysis.
  • Fig. S10. Candidate Plk targets.
  • Fig. S11. In vitro peptide kinase motif assay.
  • Fig. S12. Regulation of Aurora A by Plk.
  • Fig. S13. Prediction of phosphorylation sites in structured and ordered regions of proteins.
  • Fig. S14. Evolutionary motif conservation.
  • Fig. S15. T loop sequence alignments.
  • Fig. S16. STRING and MCODE analysis of proteins predicted to be phosphorylated by Aurora A, Aurora B, or Plk.
  • Fig. S17. Western blot and flow cytometry analysis of nocodazole-arrested HeLa cells and HeLa cells collected by mitotic shake-off after thymidine release.
  • Fig. S18. Reciprocal plots of mass spectrometry results from in vitro kinase reactions and cellular proteomics analysis.
  • Fig. S19. The effect of NuMA phosphorylation on NuMA localization.
  • Details Regarding Data Availability
  • Description for Tables S1 to S6
  • References

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Technical Details

Format: Adobe Acrobat PDF

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Other Supplementary Material for this manuscript includes the following:

  • Table S1 (Microsoft Excel format). All ModSites and representative peptides.
  • Table S2 (Microsoft Excel format). ModSites assigned to the Aurora kinase substrate cluster.
  • Table S3 (Microsoft Excel format). ModSites assigned to the Plk substrate cluster.
  • Table S4 (Microsoft Excel format). Analysis of site and motif conservation for candidate Aurora A, Aurora B, and Plk substrates across evolution.
  • Table S5 (Microsoft Excel format). ModSite assignments to Aurora A, Aurora B, Aurora ambiguous, and Plk clusters.
  • Table S6 (Microsoft Excel format). Plk1-interacting proteins.

[Download Tables S1 to S6 (Compressed)]

Technical Details

Format: Microsoft Excel. Files are packaged as a compressed archive, in *.zip format; users should download the compressed file to their machine and decompress the file on their local hard drive, using the instructions below.

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Citation: A. N. Kettenbach, D. K. Schweppe, B. K. Faherty, D. Pechenick, A. A. Pletnev, S. A. Gerber, Quantitative Phosphoproteomics Identifies Substrates and Functional Modules of Aurora and Polo-Like Kinase Activities in Mitotic Cells. Sci. Signal. 4, rs5 (2011).

© 2011 American Association for the Advancement of Science