Plexins are the cell surface receptors of semaphorins. Plexin-mediated semaphorin signaling is essential for processes such as the development of the nervous system and the cardiovascular system and regulation of immune responses and bone homeostasis. Malfunction of plexins has been associated with neurological disorder and cancer. Understanding how plexins function will pave the way for developing targeted therapeutics for fighting the associated diseases and improving neuronal regeneration after injury. The plexin intracellular region contains a GTPase Activating Protein (GAP) domain that is essential for function. In the previous period, we have identified the small GTPase Rap as the authentic substrate for the plexin GAP domain, and have determined the structural basis for how the GAP domain is activated by semaphorin-induced dimerization and how it inactivates Rap through a non-canonical catalytic mechanism. Objectives. To study additional layers of regulation mechanisms of plexins, and mutual regulation between plexins and several of their key binding partners. Research Design. Based on a new crystal structure of ours, we will first analyze the role of the inhibitory dimer in plexin regulation, a long-standing question in the fiel. Plexin signaling requires not only its RapGAP activity, but also its ability to assemble and contro the activity of a multi-protein signaling complex at the plasma membrane. Many proteins interact with plexins, but the structural basis of their actions is largely unknown. We will focus on some of the essential binding partners, address the questions how they bind plexin and exert mutual regulation with plexins.
In Aim 1 we will test an inhibitory dimer model in plexin regulation. We have determined two crystal structures of PlexinA4, which adopts a new conformation and forms a compact dimer with the GAP active site buried in the dimer interface. We propose that this dimer and the apo dimer structure of the plexin extracellular region reported previously together mediate the autoinhibited dimeric state of full-length plexin on the cell surface. Structure-based mutational analyses will be performed to test this hypothesis.
In Aim 2 we will study the basis for the opposite effects of RND1/Rac and RhoD on plexin signaling. The RhoGTPases Rac1 and RND1 interact with plexin and facilitate its binding and activation by semaphorin. In contrast, RhoD inhibits plexin signaling, although it binds plexin in the same mode with similar affinity. Our structure analyses led to a hypothesis that explains this paradox, which will be tested in this aim.
In Aim 3 we will analyze the interaction and regulation of FARPs by plexin. FARP1 and FARP2 are two related guanine nucleotide exchange factors (GEFs) that have been shown to interact directly with plexin and make essential contributing to its signaling. We will pursue a structure of the plexin/FARP complex to elucidate the basis for their interaction, and analyze how this interaction helps release the autoinhibition of FARPs.

Public Health Relevance

Plexins are important signaling proteins that control essential processes such as the development of the nervous and the cardiovascular systems and the regulation of immune responses. Malfunction of plexins is implicated in various diseases such as neurological disorders, autoimmune diseases and cancer. The goal of this study is to elucidate the regulation and signaling mechanisms of plexins, which will help fight diseases associated with malfunction of plexin signaling.

National Institute of Health (NIH)
National Institute of General Medical Sciences (NIGMS)
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Macromolecular Structure and Function C Study Section (MSFC)
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Flicker, Paula F
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University of Texas Sw Medical Center Dallas
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Pascoe, Heath G; Wang, Yuxiao; Zhang, Xuewu (2017) In Vitro Assay for the Rap GTPase-Activating Protein Activity of the Purified Cytoplasmic Domain of Plexin. Methods Mol Biol 1493:107-118
Shang, Guijun; Brautigam, Chad A; Chen, Rui et al. (2017) Structure analyses reveal a regulated oligomerization mechanism of the PlexinD1/GIPC/myosin VI complex. Elife 6:
Kuo, Yi-Chun; Zhang, Xuewu (2016) Regulation of Plexin: A Ring of Structural Twists and Turns. Neuron 91:497-9
Yoo, Sa Kan; Pascoe, Heath G; Pereira, Telmo et al. (2016) Plexins function in epithelial repair in both Drosophila and zebrafish. Nat Commun 7:12282
Marita, Morgan; Wang, Yuxiao; Kaliszewski, Megan J et al. (2015) Class A Plexins Are Organized as Preformed Inactive Dimers on the Cell Surface. Biophys J 109:1937-45
Pascoe, Heath G; Wang, Yuxiao; Zhang, Xuewu (2015) Structural mechanisms of plexin signaling. Prog Biophys Mol Biol 118:161-8
Pascoe, Heath G; Gutowski, Stephen; Chen, Hua et al. (2015) Secondary PDZ domain-binding site on class B plexins enhances the affinity for PDZ-RhoGEF. Proc Natl Acad Sci U S A 112:14852-7
Xu, Hui; He, Xiaojing; Zheng, Hui et al. (2014) Structural basis for the prion-like MAVS filaments in antiviral innate immunity. Elife 3:e01489
Wang, Yuxiao; Pascoe, Heath G; Brautigam, Chad A et al. (2013) Structural basis for activation and non-canonical catalysis of the Rap GTPase activating protein domain of plexin. Elife 2:e01279
Falta, Michael T; Pinilla, Clemencia; Mack, Douglas G et al. (2013) Identification of beryllium-dependent peptides recognized by CD4+ T cells in chronic beryllium disease. J Exp Med 210:1403-18

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