Methanothrixis widely distributed in natural and artificial anoxic environments and plays a major role in global methane emissions. It is one of only two genera that can form methane from acetate dismutation and through participation in direct interspecies electron transfer (DIET) with exoelectrogens. AlthoughMethanothrixis a significant member of many methanogenic communities, little is known about its physiology. In this study, transcriptomics helped to identify potential routes of electron transfer during DIET betweenGeobacter metallireducensandMethanothrix thermoacetophila. Additions of magnetite to cultures significantly enhanced growth by acetoclastic methanogenesis and by DIET, while granular activated carbon (GAC) amendments impaired growth. Transcriptomics suggested that the OmaF-OmbF-OmcF porin complex and the octaheme outer membranec-type cytochrome, Gmet_0930, were important for electron transport across the outer membrane ofG. metallireducensduring DIET withMx. thermoacetophila. Clear differences in the metabolism ofMx. thermoacetophilawhen grown via DIET or acetate dismutation were not apparent. However, genes coding for proteins involved in carbon fixation and a surface associated quinoprotein, SqpA, were highly expressed in all conditions. Expression of gas vesicle genes was significantly lower in DIET- than acetate-grown cells, possibly to facilitate better contact between membrane associated redox proteins during DIET. These studies reveal potential electron transfer mechanisms utilized by bothGeobacterandMethanothrixduring DIET and provide important insights into the physiology ofMethanothrixin anoxic environments.