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Purpose/Objective(s): Microbiome has been shown to affect tumorigenesis by promoting inflammation. However, the association between the upper aerodigestive microbiome and head and neck squamous cell carcinoma (HNSCC) is not well established. Hypoxia is a modifiable factor associated with poor radiation response. Our study analyzed the HNSCC tumor samples from The Cancer Genome Atlas (TCGA) to investigate the relationship between different HNSCC tumor subsites, hypoxia, and local tumor microbiome composition. Results: A total of 357 patients were included [Oral cavity (OC) = 226, Oropharynx (OPx) = 53, and Larynx/Hypopharynx (LHPx) = 78], of which 12.8%, 71.7%, and 10.3%, respectively, were HPV positive. The mean (SD) hypoxia scores were 30.18 (11.10), 24.31 (14.13), and 29.53 (12.61) in OC, OPx, and LHPx tumors, respectively, with higher values indicating greater hypoxia. The hypoxia score was significantly higher for OC tumors compared to OPx (p = 0.044) and LHPx (p = 0.002). There was no significant correlation between hypoxia and HPV status. Pseudomonas sp. in OC, Actinomyces sp. and Sulfurimonas sp. in OPx, and Filifactor, Pseudomonas and Actinomyces sp. in LHPx had the strongest association with the hypoxia score. Materials/Methods: Tumor RNAseq samples from TCGA were processed, and the R package “tmesig” was used to calculate gene expression signature, including the Buffa hypoxia (BH) score, a validated hypoxia signature using 52 hypoxia-regulated genes. Microbe relative abundances were modeled with primary tumor location and a high vs. low tertile BH score applying a gamma-distributed generalized linear regression using the “stats” package in R, with adjusted p-value < 0.05 considered significant. Conclusions: In our study, oral cavity tumors were found to be more hypoxic compared to other head and neck subsites, which could potentially contribute to their radiation resistance. For each subsite, distinct microbial populations were over-represented in hypoxic tumors in a subsite-specific manner. Further studies focusing on an association between microbiome, hypoxia, and patient outcomes are warranted.

Expression of FGFR1 controls both myoblast proliferation and differentiation. The Krüppel-like transcription factor BTEB1 demonstrates bimodal, reciprocal activity by activating the FGFR1 promoter in proliferating myoblasts and repressing the same promoter via the same DNA-binding site in differentiated myotubes., Expression of the gene encoding fibroblast growth factor receptor 1 (FGFR1) and subsequent FGFR1-mediated cell signaling controls numerous developmental and disease-related processes. The transcriptional regulation of the FGFR1 gene is central to these developmental events and serves as a molecular model for understanding transcriptional control of growth factor receptor genes. The FGFR1 promoter is activated in proliferating myoblasts via several Sp1-like binding elements. These elements display varying levels of activation potential, suggesting that unique protein-DNA complexes coordinate FGFR1 gene expression via each of these sites. The Krüppel-like factor, BTEB1/KLF9, was expressed in both proliferating myoblasts and differentiated myotubes in vitro. The BTEB1 protein was nuclear-localized in both cell types. BTEB1 activated the FGFR1 promoter via interaction with the Sp1-like binding site located at −59 bp within the FGFR1 promoter. FGFR1 gene expression is down-regulated during myogenic differentiation, and FGFR1 promoter activity is correspondingly reduced. This reduction in FGFR1 promoter activity was attributable to BTEB1 interaction with the same Sp1-like binding site located at −59 bp in the FGFR1 promoter. Therefore, BTEB1 is capable of functioning as a transcriptional activator and repressor of the same promoter via the same DNA-binding element and demonstrates a novel, bimodal role of BTEB1 during myogenesis.

Skeletal muscle development is partly characterized by myoblast proliferation and subsequent differentiation into postmitotic muscle fibers. Developmental regulation of expression of the fibroblast growth factor receptor 1 (FGFR1) gene is required for normal myoblast proliferation and muscle formation. As a result, FGFR1 promoter activity is controlled by multiple transcriptional regulatory proteins during both proliferation and differentiation of myogenic cells. The transcription factor AP-2 alpha is present in nuclei of skeletal muscle cells and suppresses myoblast proliferation in vitro. Since FGFR1 gene expression is tightly linked to myoblast proliferation versus differentiation, the FGFR1 promoter was examined for candidate AP-2 alpha binding sites. Mutagenesis studies indicated that a candidate binding site located at -1035 bp functioned as a repressor cis-regulatory element. Furthermore, mutation of this site alleviated AP-2 alpha-mediated repression of FGFR1 promoter activity. Chromatin immunoprecipitation studies demonstrated that AP-2 alpha interacted with the FGFR1 promoter in both proliferating myoblasts and differentiated myotubes. In total, these results indicate that AP-2 alpha is a transcriptional repressor of FGFR1 gene expression during skeletal myogenesis.