Tan Tracy C, Shen Yusheng, Stine Lily B, Mitchell Barbara, Okada Kyoko, McKenney Richard J, Ori-McKenney Kassandra M
Department of Molecular and Cellular Biology, University of California, Davis, California, USA.
Department of Molecular and Cellular Biology, University of California, Davis, California, USA.
J Biol Chem. 2024 Nov;300(11):107792. doi: 10.1016/j.jbc.2024.107792. Epub 2024 Sep 19.
Microtubule-associated protein, MAP1B, is crucial for neuronal morphogenesis and disruptions in MAP1B function are correlated with neurodevelopmental disorders. MAP1B encodes a single polypeptide that is processed into discrete proteins, a heavy chain (HC) and a light chain (LC); however, it is unclear if these two chains operate individually or as a complex within the cell. In vivo studies have characterized the contribution of MAP1B HC and LC to microtubule and actin-based processes, but their molecular mechanisms of action are unknown. Using in vitro reconstitution with purified proteins, we dissect the biophysical properties of the HC and LC and uncover distinct binding behaviors and functional roles for these MAPs. Our biochemical assays indicate that MAP1B HC and LC do not form a constitutive complex, supporting the hypothesis that these proteins operate independently within cells. Both HC and LC inhibit the microtubule motors, kinesin-3, kinesin-4, and dynein, and differentially affect the severing activity of spastin. Notably, MAP1B LC binds to actin filaments in vitro and can simultaneously bind and cross-link actin filaments and microtubules, a function not observed for MAP1B HC. Phosphorylation of MAP1B HC by dual-specificity, tyrosine phosphorylation-regulated kinase 1a negatively regulates its actin-binding activity without significantly affecting its microtubule-binding capacity, suggesting a dynamic contribution of MAP1B HC in cytoskeletal organization. Overall, our study provides new insights into the distinct functional properties of MAP1B HC and LC, underscoring their roles in coordinating cytoskeletal networks during neuronal development.
微管相关蛋白MAP1B对神经元形态发生至关重要，MAP1B功能的破坏与神经发育障碍相关。MAP1B编码一种单一多肽，该多肽被加工成离散的蛋白质，即重链（HC）和轻链（LC）；然而，尚不清楚这两条链在细胞内是单独发挥作用还是作为一个复合体发挥作用。体内研究已经描述了MAP1B HC和LC对基于微管和肌动蛋白的过程的贡献，但其分子作用机制尚不清楚。我们使用纯化蛋白进行体外重组，剖析了HC和LC的生物物理特性，并揭示了这些微管相关蛋白（MAPs）不同的结合行为和功能作用。我们的生化分析表明，MAP1B HC和LC不会形成组成型复合体，这支持了这些蛋白在细胞内独立发挥作用的假设。HC和LC均抑制微管运动蛋白驱动蛋白-3、驱动蛋白-4和动力蛋白，并对痉挛素的切断活性产生不同影响。值得注意的是，MAP1B LC在体外与肌动蛋白丝结合，并且可以同时结合和交联肌动蛋白丝和微管，而MAP1B HC未观察到这种功能。双特异性酪氨酸磷酸化调节激酶1a对MAP1B HC的磷酸化负调节其肌动蛋白结合活性，而不会显著影响其微管结合能力，这表明MAP1B HC在细胞骨架组织中具有动态作用。总体而言，我们的研究为MAP1B HC和LC的不同功能特性提供了新的见解，强调了它们在神经元发育过程中协调细胞骨架网络的作用。
