TY - JOUR
T1 - Structural basis for tubulin recognition by cytoplasmic linker protein 170 and its autoinhibition
AU - Mishima, Masaki
AU - Maesaki, Ryoko
AU - Kasa, Miyuki
AU - Watanabe, Takashi
AU - Fukata, Masaki
AU - Kaibuchi, Kozo
AU - Hakoshima, Toshio
PY - 2007/6/19
Y1 - 2007/6/19
N2 - Cytoplasmic linker protein 170 (CLIP-170) is a prototype of the plus end-tracking proteins that regulate microtubule dynamics, but it is obscure how CLIP-170 recognizes the microtubule plus end and contributes to polymerization rescue. Crystallographic, NMR, and mutation studies of two tandem cytoskeleton-associated protein glycine-rich (CAP-Gly) domains of CLIP-170, CAP-Gly-1 and CAP-Gly-2, revealed positively charged basic grooves of both CAP-Gly domains for tubulin binding, whereas the CAP-Gly-2 domain possesses a more basic groove and directly binds the EExEEY/F motif of the C-terminal acidic-tail ends of α-tubulin. Notably, the p150Glued CAP-Gly domain that is furnished with a less positively charged surface only weakly interacts with the α-tubulin acidic tail. Mutation studies showed that this acidic sextette motif is the minimum region for CAP-Gly binding. The C-terminal zinc knuckle domains of CLIP-170 bind the basic groove to inhibit the binding to the acidic tails. These results provide a structural basis for the proposed CLIP-170 copolymerization with tubulin on the microtubule plus end. CLIP-170 strongly binds the acidic tails of EB1 as well as those of α-tubulins, indicating that EB1 localized at the plus end contributes to CLIP-170 recruitment to the plus end. We suggest that CLIP-170 stimulates microtubule polymerization and/or nucleation by neutralizing the negative charges of tubulins with the highly positive charges of the CLIP-170 CAP-Gly domains. Once CLIP-170 binds microtubule, the released zinc knuckle domain may serve to recruit dynein to the plus end by interacting with p150Glued and LIS1. Thus, our structures provide the structural basis for the specific dynein loading on the microtubule plus end.
AB - Cytoplasmic linker protein 170 (CLIP-170) is a prototype of the plus end-tracking proteins that regulate microtubule dynamics, but it is obscure how CLIP-170 recognizes the microtubule plus end and contributes to polymerization rescue. Crystallographic, NMR, and mutation studies of two tandem cytoskeleton-associated protein glycine-rich (CAP-Gly) domains of CLIP-170, CAP-Gly-1 and CAP-Gly-2, revealed positively charged basic grooves of both CAP-Gly domains for tubulin binding, whereas the CAP-Gly-2 domain possesses a more basic groove and directly binds the EExEEY/F motif of the C-terminal acidic-tail ends of α-tubulin. Notably, the p150Glued CAP-Gly domain that is furnished with a less positively charged surface only weakly interacts with the α-tubulin acidic tail. Mutation studies showed that this acidic sextette motif is the minimum region for CAP-Gly binding. The C-terminal zinc knuckle domains of CLIP-170 bind the basic groove to inhibit the binding to the acidic tails. These results provide a structural basis for the proposed CLIP-170 copolymerization with tubulin on the microtubule plus end. CLIP-170 strongly binds the acidic tails of EB1 as well as those of α-tubulins, indicating that EB1 localized at the plus end contributes to CLIP-170 recruitment to the plus end. We suggest that CLIP-170 stimulates microtubule polymerization and/or nucleation by neutralizing the negative charges of tubulins with the highly positive charges of the CLIP-170 CAP-Gly domains. Once CLIP-170 binds microtubule, the released zinc knuckle domain may serve to recruit dynein to the plus end by interacting with p150Glued and LIS1. Thus, our structures provide the structural basis for the specific dynein loading on the microtubule plus end.
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U2 - 10.1073/pnas.0703876104
DO - 10.1073/pnas.0703876104
M3 - Article
C2 - 17563362
AN - SCOPUS:34547532472
SN - 0027-8424
VL - 104
SP - 10346
EP - 10351
JO - Proceedings of the National Academy of Sciences of the United States of America
JF - Proceedings of the National Academy of Sciences of the United States of America
IS - 25
ER -