Here we report the identification of a novel gene, MHY1
, and initial characterization of its product, Mhy1p, involved in the yeast-to-hypha transition of the dimorphic yeast Y. lipolytica
. Although the exact role of Mhy1p in the dimorphic transition remains undetermined, several features of Mhy1p suggest that it may function as a transcription factor. The most striking of these is the presence near its carboxyl terminus of two C2
-type zinc fingers, which are strongly homologous to zinc fingers found in the proteins encoded by the S. cerevisiae
and the open reading frame of unknown function, YER130C. Msn2p and Msn4p are transcriptional activators of the multistress response in S. cerevisiae
. They act via upstream STREs, which have the consensus core sequence AGGGG (or CCCCT) and which are able to mediate transcription induced by a broad range of environmental and physiological conditions, thereby enabling cells to develop tolerance to different forms of stress (cross-protection) (15
). Msn2p and Msn4p have been found to be constitutively synthesized during growth on glucose (9
), and they are activated by their translocation from the cytosol into the nucleus in response to stress conditions, such as heat shock, carbon source starvation, osmotic stress, and the presence of ethanol or sorbate (13
). Interestingly, conditions promoting the yeast-to-hypha transition led to a redistribution of Mhy1p from the cytosol, followed by its concentration in the nuclei of cells undergoing dimorphic transition.
Like Msn2p and Msn4p, Mhy1p specifically recognizes, and binds to, sequences containing the AGGGG pentanucleotide, strongly suggestive of a role for Mhy1p in the transcriptional regulation of genes containing this sequence in their promoter regions. Database analysis has revealed that most promoters of Y. lipolytica
genes contain AGGGG sequences, but they are particularly abundant in the genes HOY1
(six copies) and ICL1
(five copies). HOY1
has been shown to be directly involved in the yeast-to-hypha transition (38
), while ICL1
encodes one of the two main enzymes of the glyoxylate pathway, a strongly regulated anaplerotic cycle that in Y. lipolytica
is under the control of GPR1
, a gene also implicated in the dimorphic transition (3
). We are currently conducting experiments aimed at determining whether Mhy1p is a transcription factor and, if it is, whether it modulates gene expression through its interaction with STREs.
expression is dramatically increased during the yeast-to-hypha transition. Surprisingly, MHY1
mRNA levels were significantly decreased under conditions that otherwise would activate Msn2p- and Msn4p-mediated expression of stress-responsive genes in S. cerevisiae
, i.e., carbon source starvation and osmotic and oxidative shock. However, transcription of MHY1
was unaffected by thermal stress. It has been suggested that heat shock, and not starvation, may act synergistically with N
-acetylglucosamine to achieve full induction of mycelial growth (14
). Our results are in agreement with this hypothesis. Overall, analysis of the upstream region of MHY1
suggests a rather complex regulation of expression, involving feedback regulatory loops with possible connections to nitrogen starvation and stationary-phase maintenance. We are currently investigating whether these putative regulatory elements are functional and searching for proteins that can recognize these sequences.
In closing, it is important to point out that the study of stress response in Y. lipolytica is still incipient. Isolation of stress-responsive genes, and knowledge of the conditions they regulate, will be important for determining possible links between stress response and filamentous growth in dimorphic yeasts, two phenomena with large implications for the development of virulence by fungal pathogens.