Modulation of microtuble dynamics by the microtubule-associated protein MAP1a

Abstract

Structural microtubule-associated proteins (MAPs) are capable of interacting with tubulin dimers to regulate the various dynamic stages of microtubules. MAP1a is predominant in the neuronal cell body, axons and dendrites of mature neurons. MAP1 a has been shown to bind microtubules to promote microtubule assembly in vitro. The MAP1a heavy chain molecule is associated with three light chains. The heavy chain, and all three light chains appear to associate with microtubules independent of each other. The purpose of this project was to measure the impact of myc-tagged MAP1 a fragments and myc-tagged light chains associated with MAP1a on microtubule dynamic phases in vivo. Cells from an epithelial kidney cell line (LLCPK1) that had been permanently transfected with human GFP-alpha tubulin were transiently transfected with myc tagged MAP1a heavy and light chain fragments. Cells expressing MAP1a and light chain fragments were used to make direct observations of microtubule dynamics in living cells using fluorescence microscopy. Microtubule ends were photographed at 4-second intervals using a digital camera over a 2-minute duration. All truncated MAP1 a heavy chain fragments that contained the microtubule-binding domain were shown to associate with microtubules. MAP1a fragments containing portions of the projection domain promoted growth and stability of microtubules. Truncated fragments containing different regions of the projection domain of MAP1a demonstrated variations in their impact on microtubule dynamic events by promoting growth or inhibition of shortening phases. Similar to full length MAP1a, LC3 also appeared to promote microtubule growth and stability. Results from the present study suggest that MAP1a and LC3 promote slow, stable growth of microtubules. This type of growth may be important in the maintenance and restructuring of adult neurons.