CRN2 is an actin filament binding protein involved in the regulation of various cellular processes including cell migration and invasion. CRN2 has been implicated in the malignant progression of different types of human cancer. We used CRN2 knock-out mice for analyses as well as for crossbreeding with a Tp53/Pten knock-out glioblastoma mouse model. CRN2 knock-out mice were subjected to a phenotyping screen at the German Mouse Clinic. Murine glioblastoma tissue specimens as well as cultured murine brain slices and glioblastoma cell lines were investigated by immunohistochemistry, immunofluorescence, and cell biological experiments. Protein interactions were studied by immunoprecipitation, pull-down, and enzyme activity assays. CRN2 knock-out mice displayed neurological and behavioural alterations, e.g. reduced hearing sensitivity, reduced acoustic startle response, hypoactivity, and less frequent urination. While glioblastoma mice with or without the additional CRN2 knock-out allele exhibited no significant difference in their survival rates, the increased levels of CRN2 in transplanted glioblastoma cells caused a higher tumour cell encasement of murine brain slice capillaries. We identified two important factors of the tumour microenvironment, the tissue inhibitor of matrix metalloproteinase 4 (TIMP4) and the matrix metalloproteinase 14 (MMP14, synonym: MT1-MMP), as novel binding partners of CRN2. All three proteins mutually interacted and co-localised at the front of lamellipodia, and CRN2 was newly detected in exosomes. On the functional level, we demonstrate that CRN2 increased the secretion of TIMP4 as well as the catalytic activity of MMP14. Our results imply that CRN2 represents a pro-invasive effector within the tumour cell microenvironment of glioblastoma multiforme.

CRN2是一种肌动蛋白丝结合蛋白，参与各种细胞过程的调控，包括细胞迁移和侵袭。CRN2与不同类型的人类癌症的恶性进展有关。我们使用CRN2基因敲除小鼠进行分析以及与Tp53 / Pten基因敲除胶质母细胞瘤小鼠模型的杂交。在德国小鼠诊所对CRN2剔除小鼠进行了表型筛选。通过免疫组织化学，免疫荧光和细胞生物学实验研究了小鼠胶质母细胞瘤组织标本以及培养的鼠脑切片和胶质母细胞瘤细胞系。通过免疫沉淀，下拉法和酶活性试验研究了蛋白质相互作用。CRN2基因敲除小鼠表现出神经和行为改变，例如听力下降，减少听觉惊吓反应，机能减退和排尿次数减少。尽管有或没有附加CRN2基因敲除等位基因的成胶质细胞瘤小鼠的存活率均无显着差异，但移植的成胶质细胞瘤细胞中CRN2水平的升高却引起了鼠脑切片毛细血管的更高的肿瘤细胞包埋率。我们确定了肿瘤微环境的两个重要因素，即基质金属蛋白酶4（TIMP4）和基质金属蛋白酶14（MMP14，同义词：MT1-MMP）的组织抑制剂，作为CRN2的新型结合伴侣。这三种蛋白质相互相互作用并共定位于片状脂蛋白的前端，并且在外泌体中新发现了CRN2。在功能水平上，我们证明CRN2增加了TIMP4的分泌以及MMP14的催化活性。