SUMMARY Repulsive signaling by Semaphorins and Plexins is crucial for the development and homeostasis of the nervous, immune and cardiovascular systems. Sema7A acts as both an immune and a neural Semaphorin through PlexinC1, and A39R is a Sema7A mimic secreted by poxviruses. We report the structures of Sema7A and A39R complexed with the Semaphorin-binding module of PlexinC1. Both structures show two PlexinC1 molecules symmetrically bridged by Semaphorin dimers, in which the Semaphorin and PlexinC1 β-propellers interact in an edge-on, orthogonal orientation. Both binding interfaces are dominated by the insertion of the Semaphorin’s 4c-4d loop into a deep groove in blade 3 of the PlexinC1 propeller. A39R appears to achieve Sema7A mimicry by preserving key Plexin-binding determinants seen in the mammalian Sema7A complex, that have evolved to achieve higher affinity binding to the host-derived PlexinC1. The complex structures support a conserved Semaphorin-Plexin recognition mode, and suggest Plexins are activated by dimerization.

Carcinoma cell motility and invasion are prerequisites for tumor cell metastasis, which requires regulation of the actin cytoskeleton. Cortactin is an actin-related protein 2/3 (Arp2/3) complex-activating and filamentous (F)-actin-binding protein that is implicated in tumor cell motility and metastasis, partially by its ability to become tyrosine phosphorylated. Cortactin is encoded by the CTTN gene and maps to chromosome 11q13, a region amplified in many carcinomas, including head and neck squamous cell carcinoma (HNSCC). CTTN gene amplification is associated with lymph node metastasis and poor patient outcome, and cortactin overexpression enhances motility in tumor cells lacking 11q13 amplification. However, a direct link between increased motility and invasion has not been reported in tumor cells with chromosome 11q13 amplification and cortactin overexpression. In this study, we have examined the relationship between CTTN amplification and tumor cell motility in HNSCC. In 11 of 39 (28%) HNSCC cases, cortactin overexpression determined by immunohistochemistry correlates with lymph node metastasis and CTTN gene amplification. HNSCC cells containing cortactin gene amplification and protein overexpression display increased binding and activation of Arp2/3 complex, and were more motile and invasive than HNSCC cells lacking CTTN amplification. Down-regulation of cortactin expression in CTTN-amplified HNSCC cells by small interfering RNA impairs HNSCC motility and invasion. Treatment of HNSCC cells with the epidermal growth factor receptor inhibitor gefitinib inhibits HNSCC motility and down-regulates cortactin tyrosine phosphorylation. These data suggest that cortactin may be a valid prognostic and therapeutic marker for invasive and metastatic HNSCC and other carcinomas with 11q13 amplification. (Cancer Res 2006; 66(16): 8017-25)

The heterophilic synaptic adhesion molecules, neuroligins and neurexins, are essential for establishing and maintaining neuronal circuits by modulating the formation and maturation of synapses. The neuroligin-neurexin adhesion is Ca 2+ -dependent and regulated by alternative splicing. We report a 2.4Å structure of the complex between the mouse neuroligin-1 (NL1) cholinesterase-like domain and the mouse neurexin 1β (NX1β) LNS (laminins, neurexins, and sex hormone-binding globulin-like) domain. The structure reveals a delicate neuroligin-neurexin assembly mediated by a hydrophilic, Ca 2+ -mediated, and solvent-supplemented interface, rendering it capable of being modulated by alternative splicing and other regulatory factors. Thermodynamic data support a mechanism where splicing site B of NL1 acts by modulating a salt bridge at the edge of the NL1-NX1β interface. Mapping neuroligin mutations implicated in autism indicates most such mutations are structurally destabilizing, supporting deficient neuroligin biosynthesis and processing as a common cause for this brain disorder.

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In many animal cells, stimulation of cell surface receptors coupled to G proteins or tyrosine kinases mobilizes Ca2+ influx through store-operated Ca2+ release-activated Ca2+ (CRAC) channels. The ensuing Ca2+ entry regulates a wide variety of effector cell responses including transcription, motility, and proliferation. The physiological importance of CRAC channels for human health is underscored by studies indicating that mutations in CRAC channel genes produce a spectrum of devastating diseases including chronic inflammation, muscle weakness, and a severe combined immunodeficiency syndrome. Moreover, from a basic science perspective, CRAC channels exhibit a unique biophysical fingerprint characterized by exquisite Ca2+-selectivity, store-operated gating, and distinct pore properties and therefore serve as fascinating ion channels for understanding the biophysical mechanisms of ion permeation and gating. Studies in the last two decades have revealed the cellular and molecular choreography of the CRAC channel activation process, and it is now established that opening of CRAC channels is governed through direct interactions between the pore-forming Orai proteins, and the ER Ca2+ sensors, STIM1 and STIM2. In this review, we summarize the functional and structural mechanisms of CRAC channel regulation, focusing on recent advances in our understanding of the conformational and structural dynamics of CRAC channel gating.