Kellett Meghan D, Sharma Vibha, Sherlock Madeline E, Pugazhenthi Umarani, Rose Madison M, Joshi Molishree U, Dzieciatkowska Monika, Nguyen Vu, Reigan Philip, Hansen Kirk C, Kieft Jeffrey S, Schweppe Rebecca E
Division of Endocrinology, Metabolism, and Diabetes, Department of Medicine, University of Colorado, Aurora, CO, United States.
Department of Biochemistry and Molecular Genetics, University of Colorado, Aurora, CO, United States.
Front Oncol. 2025 May 19;15:1252544. doi: 10.3389/fonc.2025.1252544. eCollection 2025.
Advanced thyroid cancer, including papillary (PTC) and anaplastic thyroid cancer (ATC), are the leading causes of endocrine cancer deaths. Thus, there is a critical need to identify novel therapeutic targets to improve standard of care. Focal Adhesion Kinase (FAK) is overexpressed and phosphorylated in thyroid cancer and drives thyroid cancer growth, invasion, and metastasis. FAK is a nonreceptor tyrosine kinase that is autophosphorylated at tyrosine 397 (Y397) in response to integrin or growth factor receptor signaling, resulting in the recruitment of SRC proto-oncogene and downstream signaling pathways. FAK is predominately localized at the plasma membrane but has recently been shown to accumulate in the nucleus as well as the nucleolus to drive tumor growth. The nucleolus is a membraneless subnuclear organelle that is involved in ribosomal biogenesis through the transcription, processing, and assembly of ribosomal RNA (rRNA). The role of FAK in ribosome biogenesis is currently unknown.
Nuclear/nucleolar FAK localization and function were studied using genetic and pharmacological approaches. High resolution microscopy was used to study the subcellular localization of FAK. Functional and biochemical assays including transformation and clonogenic assays, polysome profiling, and nascent protein synthesis assays were utilized to assess cell growth and survival. Protein-protein interactions of FAK were determined using a proximity dependent biotinylation (BioID) proteomics approach.
We have found that pY397 FAK accumulates in the nucleolus of advanced thyroid cancer cells and that autophosphorylation of FAK at pY397 and FAK kinase activity are important for nucleolar accumulation of FAK. Furthermore, knockdown of nucleophosmin 1 (NPM1), an important structural component of the nucleolus, reduced pY397 FAK nucleolar accumulation. Functionally, we showed that nuclear FAK and FAK kinase activity are necessary for anchorage independent growth. We demonstrated that targeted degradation of FAK results in decreased protein synthesis with a specific decrease in the 60S ribosomal subunit. Using a BioID proteomics approach, we showed that autophosphorylated FAK interacts with a network of nucleolar proteins including nucleolar protein 56 (NOP56) which is a core small ribonucleoprotein (snoRNP) important for 60S ribosome biogenesis. Finally, we found that pY397 FAK co-localizes with NOP56 and that knockdown of NOP56 phenocopies FAK depletion.
Overall, these findings highlight a novel function for FAK in promoting ribosome biogenesis and suggest that nucleolar FAK represents a promising therapeutic target.
晚期甲状腺癌，包括乳头状甲状腺癌（PTC）和间变性甲状腺癌（ATC），是内分泌癌死亡的主要原因。因此，迫切需要确定新的治疗靶点以提高护理标准。粘着斑激酶（FAK）在甲状腺癌中过表达并磷酸化，驱动甲状腺癌的生长、侵袭和转移。FAK是一种非受体酪氨酸激酶，在整合素或生长因子受体信号传导的作用下，其酪氨酸397（Y397）会发生自磷酸化，从而导致SRC原癌基因的募集和下游信号通路的激活。FAK主要定位于质膜，但最近研究表明它也会在细胞核以及核仁中积累，从而驱动肿瘤生长。核仁是一种无膜的亚核细胞器，通过核糖体RNA（rRNA）的转录、加工和组装参与核糖体生物合成。目前尚不清楚FAK在核糖体生物合成中的作用。
使用基因和药理学方法研究核/核仁FAK的定位和功能。利用高分辨率显微镜研究FAK的亚细胞定位。采用功能和生化分析方法，包括转化和克隆形成分析、多核糖体分析和新生蛋白质合成分析，来评估细胞生长和存活情况。使用邻近依赖性生物素化（BioID）蛋白质组学方法确定FAK的蛋白质-蛋白质相互作用。
我们发现pY397 FAK在晚期甲状腺癌细胞的核仁中积累，并且FAK在pY397处的自磷酸化和FAK激酶活性对于FAK在核仁中的积累很重要。此外，核仁的重要结构成分核仁磷酸蛋白1（NPM1）的敲低减少了pY397 FAK在核仁中的积累。在功能上，我们表明核FAK和FAK激酶活性是锚定非依赖性生长所必需的。我们证明靶向降解FAK会导致蛋白质合成减少，特别是60S核糖体亚基的特异性减少。使用BioID蛋白质组学方法，我们表明自磷酸化的FAK与包括核仁蛋白56（NOP56）在内的核仁蛋白网络相互作用，NOP56是对60S核糖体生物合成很重要的核心小核糖核蛋白（snoRNP）。最后，我们发现pY397 FAK与NOP56共定位，并且NOP56的敲低模拟了FAK的缺失。
总体而言，这些发现突出了FAK在促进核糖体生物合成中的新功能，并表明核仁FAK是一个有前景的治疗靶点。
