Background: Mitochondrial defect is often observed in cancers while, in comparison with other metabolic cues, mitochondria mediated regulations in controlling tumorigenesis are less emphasized. Mitochondrial transcriptional factor A (TFAM) acts as a key regulatory factor to control mitochondrial DNA (mtDNA) replication and packing; the role of TFAM in modulating carcinogenesis, however, is controversial. Current study therefore aims to define TFAM mediated regulations in head and neck cancer (HNC) development. Methods: Multifaceted analyses in HNC cells genetically manipulated for TFAM were performed. Clinical correlations of TFAM and its downstream Electron Transport Chain (ETC) associated factors in regulating HNC progression were also examined in HNC specimens and different clinical databasesResults: At the cellular level, it was demonstrated that shRNA mediated TFAM silencing resulted in an enhanced cell proliferation, both in vitro and in vivo; in contrast, TFAM overexpression suppressed cell growth. Moreover, TFAM loss also facilitated cell migration and chemodrug resistance. At the molecular basis, TFAM mediated phenotypic changes could be resulting from metabolic reprogramming by directing HNC metabolism towards aerobic glycolysis, based on the detection of less respiratory capacity in accompany with greater extracellular acidification in response to TFAM loss. Interestingly, it was also found that TFAM loss upregulated ERK1/2 and Akt-mTORC-S6 signaling activity, revealing a potential "mitochondrion-to-cytoplasm" retrograde regulatory cue in controlling HNC malignancy. Clinical impact of TFAM and its downstream targets was further examined in clinical HNC tissues while the results showed that TFAM expression and mtDNA copy numbers were significant dropped in HNC tissues compared with their normal counterparts. By using clinical databases, HNC subjects with higher TFAM expression and less genetic alteration(s) exhibited better survival rates. Conclusion: Collectively, Current study uncovered a tumor suppressing role of TFAM and mitochondrial genome in determining HNC oncogenicity. This TFAM mediated regualtions are through intracellular metabolic reprogramming and mitochondria-to-cytoplasm cross-talk to activate oncogenic signals.