TY - JOUR
T1 - Dynamic behavior of GFP-CLIP-170 reveals fast protein turnover on microtubule plus ends
AU - Dragestein, Katharina A
AU - van Cappellen, Wiggert A
AU - van Haren, Jeffrey
AU - Tsibidis, George D
AU - Akhmanova, Anna
AU - Knoch, Tobias A
AU - Grosveld, Frank
AU - Galjart, Niels
PY - 2008/2/25
Y1 - 2008/2/25
N2 - Microtubule (MT) plus end-tracking proteins (+TIPs) specifically recognize the ends of growing MTs. +TIPs are involved in diverse cellular processes such as cell division, cell migration, and cell polarity. Although +TIP tracking is important for these processes, the mechanisms underlying plus end specificity of mammalian +TIPs are not completely understood. Cytoplasmic linker protein 170 (CLIP-170), the prototype +TIP, was proposed to bind to MT ends with high affinity, possibly by copolymerization with tubulin, and to dissociate seconds later. However, using fluorescence-based approaches, we show that two +TIPs, CLIP-170 and end-binding protein 3 (EB3), turn over rapidly on MT ends. Diffusion of CLIP-170 and EB3 appears to be rate limiting for their binding to MT plus ends. We also report that the ends of growing MTs contain a surplus of sites to which CLIP-170 binds with relatively low affinity. We propose that the observed loss of fluorescent +TIPs at plus ends does not reflect the behavior of single molecules but is a result of overall structural changes of the MT end.
AB - Microtubule (MT) plus end-tracking proteins (+TIPs) specifically recognize the ends of growing MTs. +TIPs are involved in diverse cellular processes such as cell division, cell migration, and cell polarity. Although +TIP tracking is important for these processes, the mechanisms underlying plus end specificity of mammalian +TIPs are not completely understood. Cytoplasmic linker protein 170 (CLIP-170), the prototype +TIP, was proposed to bind to MT ends with high affinity, possibly by copolymerization with tubulin, and to dissociate seconds later. However, using fluorescence-based approaches, we show that two +TIPs, CLIP-170 and end-binding protein 3 (EB3), turn over rapidly on MT ends. Diffusion of CLIP-170 and EB3 appears to be rate limiting for their binding to MT plus ends. We also report that the ends of growing MTs contain a surplus of sites to which CLIP-170 binds with relatively low affinity. We propose that the observed loss of fluorescent +TIPs at plus ends does not reflect the behavior of single molecules but is a result of overall structural changes of the MT end.
U2 - 10.1083/jcb.200707203
DO - 10.1083/jcb.200707203
M3 - Article
C2 - 18283108
SN - 0021-9525
VL - 180
SP - 729
EP - 737
JO - Journal of Cell Biology
JF - Journal of Cell Biology
IS - 4
ER -