TY - JOUR
T1 - Microtubule bundling by MAP65-1 protects against severing by inhibiting the binding of katanin
AU - Burkart, Graham M.
AU - Dixit, Ram
N1 - Funding Information:
We thank M. Bezanilla (Dartmouth College) for providing the cDNA for mRuby and the pDonor plasmid and D. Ehrhardt (Carnegie Institute for Science, Stanford University) for providing the GFP-KTN1 complemented ktn1-2 mutant. This work was supported by National Institutes of Health Grant R01 GM-114678.
Publisher Copyright:
© 2019 Burkart and Dixit.
PY - 2019/6/15
Y1 - 2019/6/15
N2 - The microtubule-severing enzyme katanin (KTN1) regulates the organization and turnover of microtubule arrays by the localized breakdown of microtubule polymers. In land plants, KTN1 activity is essential for the formation of linearly organized cortical microtubule arrays that determine the axis of cell expansion. Cell biological studies have shown that even though KTN1 binds to the sidewalls of single and bundled microtubules, severing activity is restricted to microtubule cross-over and nucleation sites, indicating that cells contain protective mechanisms to prevent indiscriminate microtubule severing. Here, we show that the microtubule-bundling protein MAP65-1 inhibits KTN1-mediated microtubule severing in vitro. Severing is inhibited at bundled microtubule segments and the severing rate of nonbundled microtubules is reduced by MAP65-1 in a concentration-dependent manner. Using various MAP65-1 mutant proteins, we demonstrate that efficient cross-linking of microtubules is crucial for this protective effect and that microtubule binding alone is not sufficient. Reduced severing due to microtubule bundling by MAP65-1 correlated to decreased binding of KTN1 to these microtubules. Taken together, our work reveals that cross-linking of microtubules by MAP65-1 confers resistance to severing by inhibiting the binding of KTN1 and identifies the structural features of MAP65-1 that are important for this activity.
AB - The microtubule-severing enzyme katanin (KTN1) regulates the organization and turnover of microtubule arrays by the localized breakdown of microtubule polymers. In land plants, KTN1 activity is essential for the formation of linearly organized cortical microtubule arrays that determine the axis of cell expansion. Cell biological studies have shown that even though KTN1 binds to the sidewalls of single and bundled microtubules, severing activity is restricted to microtubule cross-over and nucleation sites, indicating that cells contain protective mechanisms to prevent indiscriminate microtubule severing. Here, we show that the microtubule-bundling protein MAP65-1 inhibits KTN1-mediated microtubule severing in vitro. Severing is inhibited at bundled microtubule segments and the severing rate of nonbundled microtubules is reduced by MAP65-1 in a concentration-dependent manner. Using various MAP65-1 mutant proteins, we demonstrate that efficient cross-linking of microtubules is crucial for this protective effect and that microtubule binding alone is not sufficient. Reduced severing due to microtubule bundling by MAP65-1 correlated to decreased binding of KTN1 to these microtubules. Taken together, our work reveals that cross-linking of microtubules by MAP65-1 confers resistance to severing by inhibiting the binding of KTN1 and identifies the structural features of MAP65-1 that are important for this activity.
UR - http://www.scopus.com/inward/record.url?scp=85068197447&partnerID=8YFLogxK
U2 - 10.1091/mbc.E18-12-0776
DO - 10.1091/mbc.E18-12-0776
M3 - Article
C2 - 31017848
AN - SCOPUS:85068197447
SN - 1059-1524
VL - 30
SP - 1587
EP - 1597
JO - Molecular biology of the cell
JF - Molecular biology of the cell
IS - 13
ER -