Head and neck squamous cell carcinoma (HNSCC) is a malignancy with a worldwide distribution. Although intensive studies have been made, the underlying oncogenic mechanism of HNSCC requires further investigation. In this study, we examined the oncogenic role of activated Cdc42-associated kinase 1 (ACK1), an oncogenic tyrosine kinase, in regulating the proliferation of HNSCC cells and its underlying molecular mechanism. Results from immunohistochemical studies revealed that ACK1 was highly expressed in HNSCC tumors, with 77% (77/100) of tumors showing a high ACK1 immunoreactivity compared to 40% (8/20) of normal mucosa. Knockdown of ACK1 expression in HNSCC cells resulted in elevated p27 expression, reduced cell proliferation, and G1-phase cell cycle arrest. Rescue of ACK1 expression in the ACK1-knockdown cells suppressed p27 expression and restored cell proliferation. Compared to ACK1-knockdown cells, ACK1-rescued cells exhibited a restored p27 expression after MG132 treatment and showed an elevated level of ubiquitinated p27. Our data further showed that knockdown of ubiquitin ligase Skp2 resulted in elevated p27 expression. Importantly, the expression of p27(WT), p27(Y74F), or p27(Y89F) in ACK1-overexpressed 293T cells or ACK1-rescued SAS cells showed higher levels of tyrosyl-phosphorylated p27 and interaction with ACK1 or Skp2. However, the expression of p27(Y88F) mutant exhibited a relatively low phosphorylation level and barely bound with ACK1 or Skp2, showing a basal interaction as the control cells. These results suggested that ACK1 is highly expressed in HNSCC tumors and functions to promote cell proliferation by the phosphorylation and degradation of p27 in the Skp2-mediated mechanism.
Keywords: ACK1; HNSCC; Skp2; Tyrosine phosphorylation; p27.
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