Mutations mimicking growth factor–induced proliferation and motility characterize aggressive subtypes of mammary tumors. To unravel currently unknown players in these processes, we performed phosphoproteomic analysis on untransformed mammary epithelial cells (MCF10A) that were stimulated in culture with epidermal growth factor (EGF). We identified ladinin-1 (LAD1), a largely uncharacterized protein to date, as a phosphorylation-regulated mediator of the EGF-to-ERK pathway. Further experiments revealed that LAD1 mediated the proliferation and migration of mammary cells. LAD1 was transcriptionally induced, phosphorylated, and partly colocalized with actin stress fibers in response to EGF. Yeast two-hybrid, proximity ligation, and coimmunoprecipitation assays revealed that LAD1 bound to actin–cross-linking proteins called filamins. Cosedimentation analyses indicated that LAD1 played a role in actin dynamics, probably in collaboration with the scaffold protein 14-3-3σ (also called SFN). Depletion of LAD1 decreased the expression of transcripts associated with cell survival and inhibited the growth of mammary xenografts in an animal model. Furthermore, LAD1 predicts poor patient prognosis and is highly expressed in aggressive subtypes of breast cancer characterized as integrative clusters 5 and 10, which partly correspond to triple-negative and HER2-positive tumors. Thus, these findings reveal a cytoskeletal component that is critically involved in cell migration and the acquisition of oncogenic attributes in human mammary tumors.

模仿生长因子诱导的增殖和运动的突变是乳腺肿瘤侵袭性亚型的特征。为了揭示这些过程中目前未知的参与者，我们对用表皮生长因子 (EGF) 刺激的未转化乳腺上皮细胞 (MCF10A) 进行了磷酸蛋白质组学分析。我们鉴定出 ladinin-1 (LAD1)，这是一种迄今为止尚未被表征的蛋白质，是 EGF 至 ERK 途径的磷酸化调节介质。进一步的实验表明LAD1介导乳腺细胞的增殖和迁移。 LAD1 被转录诱导、磷酸化，并且响应 EGF 与肌动蛋白应激纤维部分共定位。酵母双杂交、邻近连接和免疫共沉淀分析表明，LAD1 与肌动蛋白交联蛋白（称为细丝蛋白）结合。共沉降分析表明，LAD1 在肌动蛋白动力学中发挥作用，可能与支架蛋白 14-3-3σ（也称为 SFN）协同作用。在动物模型中，LAD1 的缺失会降低与细胞存活相关的转录物的表达，并抑制乳腺异种移植物的生长。此外，LAD1 预测患者预后不良，并且在以整合簇 5 和 10 为特征的侵袭性乳腺癌亚型中高度表达，这部分对应于三阴性和 HER2 阳性肿瘤。因此，这些发现揭示了细胞骨架成分在人类乳腺肿瘤中与细胞迁移和致癌属性的获得至关重要。