Background: World Health Organization (WHO) grade 4 adult-type diffuse glioma is the most malignant primary tumor of the brain. Nucleolar protein 14 (NOP14) is recognized to contribute significantly to the assembly of small ribosomal subunits. However, the specific involvement of NOP14 in diverse cancers remains poorly understood, particularly its role in adult-type diffuse glioma, which has yet to be elucidated. Methods: A total of 20 adult-type diffuse glioma samples with varying WHO stages were collected. The protein level of NOP14 was detected using immunohistochemistry. Additionally, NOP14 expression in LN229 and U251 cell lines and collected clinical tissue samples was quantified using the Western blot technique. Furthermore, the correlation between NOP14 and clinicopathological features, survival rates, matrix and immune scores, and immune components was investigated using data from the Cancer Gene Atlas database. Results: NOP14 exhibited high expression in adult-type diffuse glioma patients, with the highest expression observed in the LN229 cell line. Moreover, elevated NOP14 expression was significantly correlated with poorer overall survival and demonstrated an association with unfavorable pathological features in a cohort of 703 glioblastoma (GBM) patients. Evidence of a connection between NOP14 and the tumor microenvironment was presented. Elevated NOP14 was linked to the infiltration of CD8{}^{+}T cell and factors related to epithelial-mesenchymal transition. In in vitro assay, NOP14 was capable of suppressing adult-type diffuse glioma cell invasion and metastasis. Conclusions: NOP14 holds great promise as a candidate biomarker for detecting prognostic, molecular, and immune signatures of adult-type diffuse glioma.