Abstract
MAP7 domain containing protein 3 (MAP7D3), a newly identified microtubule associated protein, has been shown to promote microtubule assembly and stability. Its microtubule binding region has been reported to consist of two coiled coil motifs located at the N-terminus. It possesses a MAP7 domain near the C-terminus and belongs to the microtubule associated protein 7 (MAP7) family. The MAP7 domain of MAP7 protein has been shown to bind to kinesin-1; however, the role of MAP7 domain in MAP7D3 remains unknown. Based on the bioinformatics analysis of MAP7D3, we hypothesized that the MAP7 domain of MAP7D3 may have microtubule binding activity. Indeed, we found that MAP7 domain of MAP7D3 bound to microtubules as well as enhanced the assembly of microtubules in vitro. Interestingly, a longer fragment MDCT that contained the MAP7 domain (MD) with the C-terminal tail (CT) of the protein promoted microtubule polymerization to a greater extent than MD and CT individually. MDCT stabilized microtubules against dilution induced disassembly. MDCT bound to reconstituted microtubules with an apparent dissociation constant of 3.0±0.5 µM. An immunostaining experiment showed that MDCT localized along the length of the preassembled microtubules. Competition experiments with tau indicated that MDCT shares its binding site on microtubules with tau. Further, we present evidence indicating that MDCT binds to the C-terminal tail of tubulin. In addition, MDCT could bind to tubulin in HeLa cell extract. Here, we report a microtubule binding region in the C-terminal region of MAP7D3 that may have a role in regulating microtubule assembly dynamics.
Highlights
The involvement of microtubules in various cellular events including intracellular transport of vesicles, cell differentiation, cell motility and mitosis is accredited to their dynamic behavior
MAP6 or Stable tubule only polypeptides (STOPs) are mainly expressed in adult neurons and regulate synaptic plasticity [12]. Another microtubule stabilizing protein that was isolated from HeLa cells is microtubule associated protein 7 (MAP7) which is known as E-Microtubule associated proteins (MAPs)-115 or ensconsin [9,13,14]
Characterization of MAP7 domain (MD), C-terminal tail (CT) and MDCT MD, CT and MDCT were isolated from BL21 (DE3) E. coli cells (Figure 2A)
Summary
The involvement of microtubules in various cellular events including intracellular transport of vesicles, cell differentiation, cell motility and mitosis is accredited to their dynamic behavior. MAP6 or Stable tubule only polypeptides (STOPs) are mainly expressed in adult neurons and regulate synaptic plasticity [12]. Another microtubule stabilizing protein that was isolated from HeLa cells is MAP7 which is known as E-MAP-115 or ensconsin [9,13,14]. This protein is mainly expressed in polarized cells like epithelial cells, Drosophila oocytes and Drosophila and murine neurons [13,15,16]. MAP7 has been shown to be essential for kinesin-1 driven functions [15,18,19]
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