Minimal Plus-end Tracking Unit of the Cytoplasmic Linker Protein CLIP-170

Benjamin A. Paulson,Alan J. Hunt,Kamlesh K. Gupta,Blake Charlebois,Eric S. Folker,Holly V. Goodson

doi:10.1074/jbc.m807675200

Abstract

Cytoplasmic linker protein 170 (CLIP-170) is the prototype microtubule (MT) plus-end tracking protein (+TIP) and is involved in regulating MT dynamics. A comprehensive understanding of the process by which CLIP-170 tracks MT plus ends would provide insight into its function. However, the precise molecular mechanism of CLIP-170 +TIP behavior is unknown, and many potential models have been presented. Here, by separating the two CLIP-170 CAP-Gly domains and their adjacent serine-rich regions into fragments of varied size, we have characterized the minimal plus-end tracking unit of CLIP-170 in vivo. Each CLIP-170 fragment was also characterized for its tubulin polymerization activity in vitro. We found that the two CAP-Gly domains have different activities, whereas CAP-Gly-1 appears incompetent to mediate either +TIP behavior or MT nucleation, a CLIP-170 fragment consisting of the second CAP-Gly domain and its adjacent serine-rich region can both track MT plus ends in vivo and induce tubulin polymerization in vitro. These observations complement recent work on CLIP-170 fragments, demonstrate that CAP-Gly motifs do not require dimerization for +TIP and polymerization-promoting activities, and provide insight into CLIP-170 function and mechanism.

Highlights

Microtubules (MTs)2 play essential roles in vital cellular processes, including intracellular transport, cell division, and signal transduction (1– 4)
We found that the two CAP-Gly domains have different activities, whereas CAP-Gly-1 appears incompetent to mediate either ؉TIP behavior or MT nucleation, a CLIP-170 fragment consisting of the second CAPGly domain and its adjacent serine-rich region can both track MT plus ends in vivo and induce tubulin polymerization in vitro
Because molecular events at the MT plus end govern whether the MT grows or shrinks, proteins that localize to the plus end are expected to play a significant role in the control of MT dynamics

Summary

Introduction

Microtubules (MTs)2 play essential roles in vital cellular processes, including intracellular transport, cell division, and signal transduction (1– 4). We found that the two CAP-Gly domains have different activities, whereas CAP-Gly-1 appears incompetent to mediate either ؉TIP behavior or MT nucleation, a CLIP-170 fragment consisting of the second CAPGly domain and its adjacent serine-rich region can both track MT plus ends in vivo and induce tubulin polymerization in vitro.

Results

Conclusion