NuSAP, a mitotic RanGTP target that stabilizes and cross-links microtubules.
Nucleolar and spindle-associated protein (NuSAP) was recently identified as a microtubule- and chromatin-binding protein in vertebrates that is nuclear during interphase. Small interfering RNA-mediated depletion of NuSAP resulted in aberrant spindle formation, missegregation of chromosomes, and ultimately blocked cell proliferation. We show here that NuSAP is enriched on chromatin-proximal microtubules at meiotic spindles in Xenopus oocytes. When added at higher than physiological levels to Xenopus egg extract, NuSAP induces extensive bundling of spindle microtubules and causes bundled microtubules within spindle-like structures to become longer. In vitro reconstitution experiments reveal two direct effects of NuSAP on microtubules: first, it can efficiently stabilize microtubules against depolymerization, and second, it can cross-link large numbers of microtubules into aster-like structures, thick fibers, and networks. With defined components we show that the activity of NuSAP is differentially regulated by Importin (Imp) alpha, Impbeta, and Imp7. While Impalpha and Imp7 appear to block the microtubule-stabilizing activity of NuSAP, Impbeta specifically suppresses aspects of the cross-linking activity of NuSAP. We propose that to achieve full NuSAP functionality at the spindle, all three importins must be dissociated by RanGTP. Once activated, NuSAP may aid to maintain spindle integrity by stabilizing and cross-linking microtubules around chromatin.
Ribbeck
Katharina
K
European Molecular Biology Laboratory, D-69117 Heidelberg, Germany. ribbeck@embl.de
Groen
Aaron C
AC
Santarella
Rachel
R
Bohnsack
Markus T
MT
Raemaekers
Tim
T
Köcher
Thomas
T
Gentzel
Marc
M
Görlich
Dirk
D
Wilm
Matthias
M
Carmeliet
Geert
G
Mitchison
Timothy J
TJ
Ellenberg
Jan
J
Hoenger
Andreas
A
Mattaj
Iain W
IW
D000818:Animals; D005260:Female; D005786:Gene Expression Regulation; D008540:Meiosis; D008870:Microtubules / Q000502:physiology* / Q000648:ultrastructure; D009865:Oocytes / Q000166:cytology / Q000502:physiology*; D021381:Protein Transport; D034622:RNA Interference; D008941:Spindle Apparatus / Q000502:physiology / Q000648:ultrastructure; D014404:Tubulin / Q000378:metabolism; D029867:Xenopus Proteins / Q000378:metabolism; D014982:Xenopus laevis / Q000502:physiology*; D020931:ran GTP-Binding Protein / Q000235:genetics / Q000378:metabolism*
D014404:Tubulin; D029867:Xenopus Proteins; D020931:ran GTP-Binding Protein