Regulation of microtubule destabilizing activity of Op18/stathmin downstream of Rac1.
In the leading edge of migrating cells, a subset of microtubules exhibits net growth in a Rac1- and p21-activated kinase-dependent manner. Here, we explore the possibility of whether phosphorylation and inactivation of the microtubule-destabilizing protein Op18/stathmin could be a mechanism regulating microtubule dynamics downstream of Rac1 and p21-activated kinases. We find that, in vitro, Pak1 phosphorylates Op18/stathmin specifically at serine 16 and inactivates its catastrophe promoting activity in biochemical and time lapse microscopy microtubule assembly assays. Furthermore, phosphorylation of either serine 16 or 63 is sufficient to inhibit Op18/stathmin in vitro. In cells, the microtubule-destabilizing effect of an excess of Op18/stathmin can be partially overcome by expression of constitutively active Rac1(Q61L), which is dependent on Pak activity, suggesting that the microtubule cytoskeleton can be regulated through inactivation of Op18/stathmin downstream of Rac1 and Pak in vivo. However, in vivo, Pak1 activity alone is not sufficient to phosphorylate Op18, indicating that additional pathways downstream of Rac1 are required for Op18 regulation.
Wittmann
Torsten
T
Department of Cell Biology, The Scripps Research Institute, La Jolla, California 92037, USA. twittman@scripps.edu
Bokoch
Gary M
GM
Waterman-Storer
Clare M
CM
D000595:Amino Acid Sequence; D000818:Animals; D002465:Cell Movement; D019281:Dimerization; D004591:Electrophoresis, Polyacrylamide Gel; D005982:Glutathione Transferase / Q000378:metabolism; D006801:Humans; D008868:Microtubule Proteins; D008870:Microtubules / Q000737:chemistry / Q000378:metabolism*; D008969:Molecular Sequence Data; D016366:Open Reading Frames; D010750:Phosphoproteins / Q000737:chemistry / Q000502:physiology*; D010766:Phosphorylation; D011994:Recombinant Proteins / Q000737:chemistry; D012617:Sea Urchins; D017386:Sequence Homology, Amino Acid; D012694:Serine / Q000737:chemistry; D050777:Stathmin; D013997:Time Factors; D014404:Tubulin / Q000737:chemistry; D020830:rac1 GTP-Binding Protein / Q000737:chemistry* / Q000378:metabolism
D008868:Microtubule Proteins; D010750:Phosphoproteins; D011994:Recombinant Proteins; C493995:STMN1 protein, human; D050777:Stathmin; D014404:Tubulin; D012694:Serine; D005982:Glutathione Transferase; D020830:rac1 GTP-Binding Protein