The bulk of grape juice fermentation is carried out by the yeast Saccharomyces cerevisiae, but non-Saccharomyces yeasts can modulate many sensorial aspects of the final products in ways not well understood. In this study, some of such non-conventional yeasts were screened as mixed starter cultures in a defined growth medium in both simultaneous and sequential inoculations. One strain of Starmerella bacillaris and another of Zygosaccharomyces bailii were chosen by their distinct phenotypic footprint and their ability to reduce ethanol levels at the end of fermentation. S. bacillaris losses viability strongly at the end of mixed fermentations, while Z. bailii remains viable. S. cerevisiae viability was unchanged by the presence of the other yeasts. Physiological characterization of both strains indicates that S. bacillaris behavior is overall more different from S. cerevisiae than Z. bailii. In addition, S. cerevisiae transcriptome changes to a bigger degree in the presence of S. bacillaris in comparison to mixed fermentation with Z. bailii. S. bacillaris induces the translation machinery and repress vesicular transport. Both non-Saccharomyces yeasts induce S. cerevisiae glycolytic genes, and that may be related to ethanol lowering, but some aspects of carbon-related mechanisms are specific for each strain. Z. bailii presence increases the stress-related polysaccharides trehalose and glycogen, while S. bacillaris induces gluconeogenesis genes.