History-Dependent Catastrophes Regulate Axonal Microtubule Behavior

Abstract

In Chinese hamster ovary cells, microtubules originate at the microtubule organizing center (MTOC) and grow persistently toward the cell edge, where they undergo catastrophe. In axons, microtubule dynamics must be regulated differently because microtubules grow parallel to the plasma membrane and there is no MTOC. GFP-tagged microtubule plus end tracking proteins (+TIPs) mark the ends of growing neuronal microtubules. Their fluorescent "comet-like" pattern reflects turnover of +TIP binding sites. Using GFP-tagged +TIPs and fluorescence-based segmentation and tracking tools, we show that axonal microtubules grow with a constant average velocity and that they undergo catastrophes at random positions, yet in a programmed fashion. Using protein depletion approaches, we find that the +TIPs CLIP-115 and CLIP-170 affect average microtubule growth rate and growth distance in neurons but not the duration of a microtubule growth event. In N1E-115 neuroblastoma cells, we find that EB1, the core +TIP, regulates microtubule growth rate, growth distance, and duration, consistent with in vitro data. Combined, our data suggest that CLIPs influence the axonal microtubule/tubulin ratio, whereas EB1 stimulates microtubule growth and structural transitions at microtubule ends, thereby regulating microtubule catastrophes and the turnover of +TIP binding sites.

A defect in the CLIP1 gene (CLIP-170) can cause autosomal recessive intellectual disability

“…CLIP2, a brain-specific cytoplasmic linker protein, has the same structure as CLIP1 but lacks the C-terminal metal-binding motifs. 23 Although the exact function of CLIP2 is unknown, the protein is involved in regulating MT dynamics in neurons 24 and in nervous system development and function. 16 In human, the CLIP2 gene is located in the region that is commonly deleted in Williams-Beuren syndrome (MIM 194050) patients.…”

Theoretical Analysis of Microtubule Dynamics at All Times

“…2,3,8,11,12 Another issue is that even more detailed theoretical investigations concentrate mostly on stationarystate behavior of the system. [13][14][15][16] In other words, dynamic properties of microtubules are assumed to be stationary and independent of the time.…”

Regulation of EB1/3 proteins by classical MAPs in neurons

“…26 Both EB1 and EB3 have been shown to be involved in neurite outgrowth. 29,30 Downregulation of EB1 leads to a reduction in neurite length in differentiating neuroblastoma cells in which EB1 regulates MT growth rate, growth distance and duration. 29 In Drosophila neurons, EB1 is required for uniform dendrite MT polarity and MT growth at dendrite branch points.…”

“…29,30 Downregulation of EB1 leads to a reduction in neurite length in differentiating neuroblastoma cells in which EB1 regulates MT growth rate, growth distance and duration. 29 In Drosophila neurons, EB1 is required for uniform dendrite MT polarity and MT growth at dendrite branch points. 31 EB1 is enriched in the distal axon, recruiting the dynein activator dynactin in an ordered pathway, thus leading to the initiation of retrograde transport by the motor protein dynein.…”