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Economically feasible bioethanol process from lignocellulose requires efficient fermentation by yeast of all sugars present in the hydrolysate. However, when exposed to lignocellulosic hydrolysate, <i>Saccharomyces cerevisiae</i> is challenged with a variety of inhibitors that reduce yeast viability, growth, and fermentation rate, and in addition damage cellular structures. In order to evaluate the capability of <i>S. cerevisiae</i> to adapt and respond to lignocellulosic hydrolysates, the physiological effect of cultivating yeast in the spruce hydrolysate was comprehensively studied by assessment of yeast performance in simultaneous saccharification and fermentation (SSF), measurement of furaldehyde reduction activity, assessment of conversion of phenolic compounds and genome-wide transcription analysis. The yeast cultivated in spruce hydrolysate developed a rapid adaptive response to lignocellulosic hydrolysate, which significantly improved its fermentation performance in subsequent SSF experiments. The adaptation was shown to involve the induction of NADPH-dependent aldehyde reductases and conversion of phenolic compounds during the fed-batch cultivation. These properties were correlated to the expression of several genes encoding oxidoreductases, notably <i>AAD4, ADH6, OYE2/3,</i> and YML131w. The other most significant transcriptional changes involved genes involved in transport mechanisms, such as <i>YHK8, FLR1,</i> or <i>ATR1</i>. A large set of genes were found to be associated with transcription factors (TFs) involved in stress response (Msn2p, Msn4p, Yap1p) but also cell growth and division (Gcr4p, Ste12p, Sok2p), and these TFs were most likely controlling the response at the post-transcriptional level.