FIGURE 6.

A transgenic mouse model verifies that Mettl1 regulates PI3K/Akt/mTOR activity in HNSCC. (A) UMAP of scRNA‐seq cells recovered from both Mettl1^cKO‐Ctrl and Mettl1^cKO mice labeled by cell type (left). Dot plot of selected subset‐associated genes across cell types from scRNA‐seq data (right). (B) UMAP of reclustered epithelial cells from Mettl1^cKO‐Ctrl and Mettl1^cKO mice. (C) Violin plots of the expression of the Mettl1 gene in epithelial cells from Mettl1^cKO‐Ctrl and Mettl1^cKO mice. (D) Violin plots of the GSVA results for epithelial cells revealed that PI3K/Akt activation was significantly decreased in Mettl1^cKO HNSCC samples. (E) qRT‐PCR of Pik3ca expression (K14CreER;RiboTag;Mettl1^fl/fl /K14CreER;RiboTag;Mettl1^wt/wt ) in input RNA was normalized to that of Gapdh and the ratio of IP RNA. (F) Representative images (left) of tongue lesions from Mettl1^cKI‐Ctrl and Mettl1^cKI mice by oral gavage of BKM120 (Mettl1^cKI‐Ctrl + BKM120 and Mettl1^cKI + BKM120) compared with Mettl1^cKI‐Ctrl and Mettl1^cKI mice by oral gavage of buffer agent (Mettl1^cKI‐Ctrl and Mettl1^cKI). Quantification of the number of lesions (middle) and the lesion areas (right) visible in the tongue (n = 6). (G) Representative H&E staining of lesions (left) from Mettl1^cKI‐Ctrl, Mettl1^cKI, Mettl1^cKI‐Ctrl + BKM120 and Mettl1^cKI + BKM120 mice. Quantification of histological grade of lesions in different groups (right). (H) Representative IHC staining (left) and quantification (right) of PI3K in Mettl1^cKI‐Ctrl, Mettl1^cKI, Mettl1^cKI‐Ctrl + BKM120 and Mettl1^cKI + BKM120 mice. Data are presented as the mean ± SD and analyzed by Student's t‐test or Chi‐squared Test. *, P < 0.05, **, P < 0.01, ***, P < 0.001. Abbreviations: PI3K/Akt/mTOR: phosphatidylinositol‐3‐kinase/protein kinase B/mammalian target of rapamycin; Pik3ca: phosphatidylinositol‐4,5‐bisphosphate 3‐kinase, catalytic subunit alpha; Mettl1: Methyltransferase‐like 1; IHC: immunohistochemistry; IP: immunoprecipitation; UMAP: Uniform manifold approximation and projection; GSVA: gene set variation analysis; Mettl1^cKO‐Ctrl: K14CreER;Mettl1^wt/wt mice; Mettl1^cKO: K14CreER;Mettl1^fl/fl mice; Mettl1^cKI‐Ctrl: K14Cre;Mettl1^wt/wt mice; Mettl1^cKI: K14Cre;Mettl1^KI/KI mice; SD: standard deviation