PDF(Pubmed)
Abstract：
Head and neck squamous cell carcinoma (HNSCC) is a major health concern due to its high mortality from poor treatment responses and locoregional tumor invasion into life sustaining structures in the head and neck. A deeper comprehension of HNSCC invasion mechanisms holds the potential to inform targeted therapies that may enhance patient survival. We previously reported that doublecortin like kinase 1 (DCLK1) regulates invasion of HNSCC cells. Here, we tested the hypothesis that DCLK1 regulates proteins within invadopodia to facilitate HNSCC invasion. Invadopodia are specialized subcellular protrusions secreting matrix metalloproteinases that degrade the extracellular matrix (ECM). Through a comprehensive proteome analysis comparing DCLK1 control and shDCLK1 conditions, our findings reveal that DCLK1 plays a pivotal role in regulating proteins that orchestrate cytoskeletal and ECM remodeling, contributing to cell invasion. Further, we demonstrate in TCGA datasets that DCLK1 levels correlate with increasing histological grade and lymph node metastasis. We identified higher expression of DCLK1 in the leading edge of HNSCC tissue. Knockdown of DCLK1 in HNSCC reduced the number of invadopodia, cell adhesion and colony formation. Using super resolution microscopy, we demonstrate localization of DCLK1 in invadopodia and colocalization with mature invadopodia markers TKS4, TKS5, cortactin and MT1-MMP. We carried out phosphoproteomics and validated using immunofluorescence and proximity ligation assays, the interaction between DCLK1 and motor protein KIF16B. Pharmacological inhibition or knockdown of DCLK1 reduced interaction with KIF16B, secretion of MMPs, and cell invasion. This research unveils a novel function of DCLK1 within invadopodia to regulate the trafficking of matrix degrading cargo. The work highlights the impact of targeting DCLK1 to inhibit locoregional invasion, a life-threatening attribute of HNSCC.
摘要：
头颈部鳞状细胞癌（HNSCC）是一个主要的健康问题，因为它的高死亡率来自不良的治疗反应和局部肿瘤侵入头颈部的生命维持结构。对HNSCC侵袭机制的更深入理解具有告知可能增强患者生存的靶向治疗的潜力。我们先前报道了双皮质素样激酶1（DCLK1）调节HNSCC细胞的侵袭。这里,我们检验了DCLK1调节invadopodia内的蛋白质以促进HNSCC入侵的假设。Invadopodia是分泌降解细胞外基质（ECM）的基质金属蛋白酶的特化亚细胞突起。通过比较DCLK1对照和shDCLK1条件的综合蛋白质组分析，我们的研究结果表明，DCLK1在调节细胞骨架和ECM重塑的蛋白质中起着关键作用，有助于细胞入侵。Further,我们在TCGA数据集中证明DCLK1水平与组织学分级和淋巴结转移增加相关.我们发现DCLK1在HNSCC组织的前缘有较高的表达。在HNSCC中敲除DCLK1减少了invadopodia的数量，细胞粘附和集落形成。使用超分辨率显微镜，我们证明了DCLK1在侵袭足中的定位以及与成熟的侵袭足病标记TKS4，TKS5，cortactin和MT1-MMP的共定位。我们进行了磷酸蛋白质组学，并使用免疫荧光和邻近连接试验进行了验证，DCLK1与运动蛋白KIF16B之间的相互作用。药物抑制或敲低DCLK1减少与KIF16B的相互作用，MMPs的分泌，和细胞入侵。这项研究揭示了DCLK1在invadopodia中的新功能，以调节基质降解货物的贩运。这项工作强调了靶向DCLK1抑制局部区域入侵的影响，HNSCC的威胁生命的属性。
