TY - JOUR
T1 - Interaction of spindle assembly factor TPX2 with importins-α/β inhibits protein phase separation
AU - Safari, Mohammad S.
AU - King, Matthew R.
AU - Brangwynne, Clifford P.
AU - Petry, Sabine
N1 - Publisher Copyright:
© 2021 THE AUTHORS.
PY - 2021/9/1
Y1 - 2021/9/1
N2 - The microtubule-based mitotic spindle is responsible for equally partitioning the genome during each cell division, and its assembly is executed via several microtubule nucleation pathways. Targeting Protein for XKlp2 (TPX2) stimulates the branching microtubule nucleation pathway, where new microtubules are nucleated from preexisting ones within mitotic or meiotic spindles. TPX2, like other spindle assembly factors, is sequestered by binding to nuclear importins-α/β until the onset of mitosis, yet the molecular nature of this regulation remains unclear. Here we demonstrate that TPX2 interacts with importins-α/β with nanomolar affinity in a 1:1:1 monodispersed trimer. We also identify a new nuclear localization sequence in TPX2 that contributes to its high-affinity interaction with importin-α. In addition, we establish that TPX2 interacts with importin-β via dispersed, weak interactions. We show that interactions of both importin-α and -β with TPX2 inhibit its ability to undergo phase separation, which was recently shown to enhance the kinetics of branching microtubule nucleation. In summary, our study informs how importins regulate TPX2 to facilitate spindle assembly, and provides novel insight into the functional regulation of protein phase separation.
AB - The microtubule-based mitotic spindle is responsible for equally partitioning the genome during each cell division, and its assembly is executed via several microtubule nucleation pathways. Targeting Protein for XKlp2 (TPX2) stimulates the branching microtubule nucleation pathway, where new microtubules are nucleated from preexisting ones within mitotic or meiotic spindles. TPX2, like other spindle assembly factors, is sequestered by binding to nuclear importins-α/β until the onset of mitosis, yet the molecular nature of this regulation remains unclear. Here we demonstrate that TPX2 interacts with importins-α/β with nanomolar affinity in a 1:1:1 monodispersed trimer. We also identify a new nuclear localization sequence in TPX2 that contributes to its high-affinity interaction with importin-α. In addition, we establish that TPX2 interacts with importin-β via dispersed, weak interactions. We show that interactions of both importin-α and -β with TPX2 inhibit its ability to undergo phase separation, which was recently shown to enhance the kinetics of branching microtubule nucleation. In summary, our study informs how importins regulate TPX2 to facilitate spindle assembly, and provides novel insight into the functional regulation of protein phase separation.
UR - http://www.scopus.com/inward/record.url?scp=85113822199&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=85113822199&partnerID=8YFLogxK
U2 - 10.1016/j.jbc.2021.100998
DO - 10.1016/j.jbc.2021.100998
M3 - Article
C2 - 34302807
AN - SCOPUS:85113822199
SN - 0021-9258
VL - 297
JO - Journal of Biological Chemistry
JF - Journal of Biological Chemistry
IS - 3
M1 - 10098
ER -