Localized alteration of microtubule polymerization in response to guidance cues.
Inhibition of microtubule dynamic instability prevents growth cone turning in response to guidance cues, yet specific changes in microtubule polymerization as growth cones encounter boundaries have not been investigated. In this study, we examined the rate and direction of microtubule polymerization in response to soluble nerve growth factor (NGF) and immobilized chondroitin sulfate proteoglycans (CSPGs) by expressing enhanced GFP-EB3 in rat pheochromocytoma (PC12) cells. GFP-EB3 comets were monitored in live cells using time-lapse epifluorescent microscopy. With an automated tracking system, the rate of microtubule polymerization was calculated as the frame-to-frame displacement of EB3 comets. Our results demonstrate that the rate of microtubule polymerization is increased following NGF treatment, whereas contact with CSPGs decreases microtubule polymerization rates. This reduction in microtubule polymerization rates was specifically localized to neurites in direct contact with CSPGs and not at noncontacting neurites. Additionally, we found an increase in the percentage of microtubules polymerizing in the retrograde direction in neurites at CSPG boundaries, with a concomitant decrease in the rate of retrograde microtubule polymerization. These results implicate localized changes in microtubule dynamics as an important component of the growth cone response to guidance cues.
Kelly
Terri-Ann N
TA
Developmental Neurobiology Section, Cell Biology and Physiology Center, National Heart, Lung, and Blood Institute, National Institutes of Health, Bethesda, MD 20892, USA.
Katagiri
Yasuhiro
Y
Vartanian
Keri B
KB
Kumar
Pramukta
P
Chen
Inn-Inn
II
Rosoff
William J
WJ
Urbach
Jeffery S
JS
Geller
Herbert M
HM
D000818:Animals; D002454:Cell Differentiation / Q000502:physiology; D011508:Chondroitin Sulfate Proteoglycans / Q000737:chemistry / Q000502:physiology; D003463:Cues; D020439:Growth Cones / Q000737:chemistry / Q000502:physiology*; D008870:Microtubules / Q000737:chemistry / Q000502:physiology*; D020932:Nerve Growth Factor / Q000737:chemistry / Q000502:physiology; D009434:Neural Pathways / Q000737:chemistry / Q000166:cytology / Q000196:embryology; D055495:Neurogenesis / Q000502:physiology; D016716:PC12 Cells; D058105:Polymerization; D051381:Rats; D015398:Signal Transduction / Q000502:physiology
D011508:Chondroitin Sulfate Proteoglycans; D020932:Nerve Growth Factor
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