Yadav Saroj, Verma Paul J, Panda Dulal
IITB-Monash Research Academy, Indian Institute of Technology Bombay, Mumbai, Maharashtra, India.
Centre for Reproduction and Development, Monash Institute of Medical Research, Monash University, Clayton, Victoria, Australia.
PLoS One. 2014 Jun 13;9(6):e99539. doi: 10.1371/journal.pone.0099539. eCollection 2014.
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.
含MAP7结构域蛋白3（MAP7D3）是一种新发现的微管相关蛋白，已被证明可促进微管组装和稳定性。据报道，其微管结合区域由位于N端的两个卷曲螺旋基序组成。它在C端附近具有一个MAP7结构域，属于微管相关蛋白7（MAP7）家族。已证明MAP7蛋白的MAP7结构域可与驱动蛋白-1结合；然而，MAP7结构域在MAP7D3中的作用仍不清楚。基于对MAP7D3的生物信息学分析，我们推测MAP7D3的MAP7结构域可能具有微管结合活性。事实上，我们发现MAP7D3的MAP7结构域可与微管结合，并在体外增强微管的组装。有趣的是，包含MAP7结构域（MD）和该蛋白C端尾巴（CT）的较长片段MDCT比单独的MD和CT更能促进微管聚合。MDCT可稳定微管，防止因稀释而解聚。MDCT与重组微管结合，表观解离常数为3.0±0.5μM。免疫染色实验表明，MDCT沿预组装微管的长度定位。与tau的竞争实验表明，MDCT与tau在微管上共享结合位点。此外，我们提供的证据表明MDCT与微管蛋白的C端尾巴结合。此外，MDCT可与HeLa细胞提取物中的微管蛋白结合。在此，我们报告了MAP7D3 C端区域的一个微管结合区域，该区域可能在调节微管组装动力学中发挥作用。
