The yeast Saccharomyces cerevisiae contains a flavohemoglobin, encoded by the gene YHB1, whose function is unclear. Previous reports presented evidence that its maximal expression requires disruption of mitochondrial respiration and that it plays a role in the response to oxidative stress. We have studied the expression of YHB1 in respiratory deficient cells and in cells exposed to various compounds causing oxidative stress. Several different strains and approaches (spectroscopic detection of the oxygenated form of Yhb1p, beta-galactosidase activity of a YHB1-lacZ fusion, and Northern blot analysis) were used to demonstrate that YHB1 expression and Yhb1p production are not increased by respiration deficiency. YHB1 expression was unchanged in cells challenged with antimycin A or menadione, while it decreased in cells exposed to H2O2, diamide, dithiothreitol, and Cu2+. Transcription of YHB1 is not under the control of the transcriptional factor Yap1p. These results do not support a participation of YHB1 in the genetic response to oxidative stress. We also analyzed the growth phenotypes associated with altered Yhb1p production using YHB1-deleted strains and strains that greatly overproduced Yhb1p. Yhb1p appears to protect cells against the damage caused by Cu2+ and dithiothreitol, while sensitizing them to H2O2. Yhb1p overproduction in a glucose-6-phosphate dehydrogenase-deficient mutant decreased its growth rate. These data indicate that there is a complex relationship(s) between Yhb1p function(s) and cell defense reactions against various stresses.