Semaphorin activates Hippo signaling to control cell growth

Semaphorins play critical roles in the tumorigenesis of various organs. Originally identified as neuronal axonal guidance molecules and developmental regulators of the nervous and the vascular systems, class 3 Semaphorins (SEMA3s) act through their receptors Plexin As (PLXNAs) and co-receptors Neuropilins (NRPs) to control various processes such as axon growth cone directionality, cell cycle, and angiogenesis. Deletion or silencing of SEMA3B and SEMA3F genes has been observed in lung cancer, metastatic breast cancer, renal carcinoma, and many other malignancies; however, the downstream effectors of SEMA3 in tumorigenesis are still unclear. Here we show that the Hippo pathway is a key mediator of SEMA3's tumor suppressive function. SEMA3 activates the Hippo pathway to control gene expression and inhibit cell growth. Restoration of SEMA3B expression in lung cancer cells that harbor SEMA3B deletion or silencing suppressed anchorage-independent growth and xenograft formation in a Hippo pathway-dependent manner. Mechanistically, PLXNA promotes the interaction and activation of p190RhoGAPs (ARHGAP5 and ARHGAP35) by the RND GTPase. Activated ARHGAP5/35 in turn act through RhoA and the Hippo kinase cascade to phosphorylate and inactivate YAP and TAZ, the transcriptional co-activators and key effectors of the Hippo pathway. Cancer-associated mutations of RND or ARHGAP5/35 compromised cellular responses to SEMA3, as indicated by YAP phosphorylation and cell growth inhibition. Our study defines a new role of the Hippo pathway in SEMA3 signaling as well as a mechanism for the tumor suppressive function of Semaphorins. Overall design: The overall goal is to examine expression changes in response to class 3 Semaphorins (SEMA3s). mRNA profiles of wild type (WT), LATS1/2 dKO (KO) upon SEMA3s treatment for U87-MG cells were generated using deep sequencing with three biological replicates.

脑信号蛋白（Semaphorins）在多种器官的肿瘤发生过程中发挥关键作用。这类蛋白最初被鉴定为神经元轴突导向分子，以及神经系统与血管系统的发育调控因子；3类脑信号蛋白（SEMA3s）通过其受体丛蛋白A家族（PLXNAs）与共受体神经纤毛蛋白（NRPs）发挥功能，调控轴突生长锥导向、细胞周期、血管生成等诸多生物学过程。研究人员已在肺癌、转移性乳腺癌、肾癌及多种其他恶性肿瘤中，检测到SEMA3B与SEMA3F基因的缺失或沉默；但目前学界对于SEMA3在肿瘤发生过程中的下游效应分子仍不明确。本研究证实，Hippo通路（Hippo pathway）是SEMA3发挥肿瘤抑制功能的关键介导因子。SEMA3可通过激活Hippo通路调控基因表达，并抑制细胞增殖。在存在SEMA3B缺失或沉默的肺癌细胞中恢复SEMA3B的表达，可通过依赖于Hippo通路的方式，抑制其非锚定依赖性生长与异种移植瘤形成。从分子机制来看，PLXNA可促进RND GTP酶（RND GTPase）介导p190RhoGAPs（ARHGAP5与ARHGAP35）的相互作用与激活。激活后的ARHGAP5/35进而通过RhoA与Hippo激酶级联反应，使作为Hippo通路转录共激活因子与关键效应分子的YAP与TAZ发生磷酸化并失活。RND或ARHGAP5/35的癌症相关突变会削弱细胞对SEMA3的应答反应，这一点可通过YAP磷酸化水平下调与细胞增殖抑制效应得以验证。本研究不仅明确了Hippo通路在SEMA3信号传导中的全新功能，同时阐明了脑信号蛋白发挥肿瘤抑制作用的分子机制。整体实验设计：本研究的核心目标是探究细胞在受到3类脑信号蛋白（SEMA3s）刺激后的基因表达变化。我们对U87-MG细胞分别设置野生型（WT）与LATS1/2双敲除（KO）两组，经SEMA3s刺激后，通过深度测序获取其mRNA表达谱，每组设置3次生物学重复。