Stress em Levedura

Interesse da Investigação

Oxidative and cold stress, signalling, transcription.

Membros do Grupo

João Coelho	External Masters Student
	Tel: 21 446 4610
Dora Pinto	Trainee
	Tel: 21 440 7900

Projecto de Investigação

Transactivator Yap1 - crossroad of cold and oxidative stress signaling pathways in Saccharomyces cerevisiae

Eukaryotic cells respond to both stimuli, like oxidative stress and temperature fluctuations, by activating cascades which are diverse but with discrete specificity. How does the cell keep track of multiple cascades? What are the cross-talks between such cascades and how are they regulated? The Yap family of bZIP transcriptional factors in Saccharomyces cerevisiae contains members which are central players in cellular responses to stress challenges as: Yap1 in oxidative and cold signals, Yap4 in response to compounds affecting cytoskeleton as well as in cold-sensitivity of yap1-deleted cells, and Yap8 in arsenite response. Although it has been previously suggested that each Yap family member plays distinct biological function, the involvement of Yap proteins in different phenomena emphasizes putative overlaping among the respectives signaling cascades. From this point of view, Yap constitutes an excelent tool to address the cross-talk of signalling pathways and, in particular, it will also provides clues on the cellular mechanisms for adaptation to low temperatures. Our major aim has been focussed on understanding the crossroad of cold and oxidative signaling pathways mediated by Yaps. The involvement of Yaps, as the target regulatory proteins, in both phenomena has been ascertained by addressing (a) the specific role of Yap1 under cold signals as well as (b) the specific role of YAP4 under equivalent conditions. Understanding the specific role of Yap1 under cold signals has been primarily addressed by determining the type of transcriptional activity of this regulator and by studying cellular transcriptional pattern putatively mediated by this factor in those conditions. The specific role of YAP4 under same conditions was approached by determining the type of transcriptional activity of this regulator as well as by developing a genetic screening to identify genes essential to the cold-phenotype of yap4 cells. This loss-of-function genetic screening has generated few mutants that revert yap4 cold-sensitivity. The identification of such mutations by complementation assays are being undertaken.

Funding

POCTI/BCI/37862/2001
Transactivator Yap1 – crossroad of cold and oxidative stress signaling pathways in Saccharomyces cerevisiae.

Projecto de Investigação

Mechanism of activation by Yap1:-signaling through RNA polymerase II basal machinery.

Yeast Saccharomyces cerevisiae as well as other fungi contains a set of non classical AP-1 factors, the Yaps, that are similar in structural motifs yet distinct in their amino acid sequences. Yaps are described as key proteins in cellular response to specific stress signals. Transactivators, like Yap1, stimulate gene expression by binding regulatory cis-elements, contacting directly or indirectly components of the RNA polymerase II basal machinery (GTFs as TFIIA, TFIIB, components of TFIID) or the Mediator, as well as by recruiting the nucleosome-remodeling complexes. In order to understand the specific signaling downstream of Yap1, we are addressing the role of GTFs as specific targets under oxidative conditions generated by hydrogen peroxide. In this context, we have selected a GTF as primary target for Yap1 activity, and we are studying the mechanism that regulates its gene expression as well as the biological relevance of such regulation under oxidative contexts. To further explore the relevance of the GTF to Yap1-mediated transcription, gtf mutants with impaired oxidative stress response (but with normal physiological behaviour) have been generated and the mutations are currently being identified. Further characterization will address the relevance of such mutation on Yap1 activity.

Projecto de Investigação

Molecular mechanisms bridging protein folding and transcription activation.

Funding

PDCT/BIA-BCM/55501/2004
Molecular mechanisms bridging protein folding and transcription activation

Publicações

Faria, J.P., Fernandes, L. (2006). Protection against oxidative stress through SUA7/TFIIB regulation in Saccharomyces cerevisiae. Free Radic. Biol. Med. (in press)

Dionisio, F., Conceição, I.C., Marques A.C.R., Fernandes, L. and Gordo, I. (2005). The evolution of a conjugative plasmid and its ability to increase bacterial fitness Biol. Lett. 1 :250-252

Cyrne, L., Martins, L., Fernandes, L., Marinho, H.S. (2003). Regulation of antioxidant enzymes gene expression in the yeast Saccharomyces cerevisiae during stationary phase Free Radic. Biol. Med. 34 :385-393

Fernandes, L., Rodrigues-Pousada, C., Struhl, K. (1997). Yap, a novel family of eight bZIP proteins in Saccharomyces cerevisiae with distinct biological functions Mol. Cell. Biol. 17 :6982-6993

Bossier, P., Fernandes, L., Vilela, C., Rodrigues-Pousada, C. (1994). The yeast YKL714 gene situated on the left arm of chromosome XI codes for an homolog of the human ALD protein Yeast 10 :681-686

Dujon, B. (Ö), Fernandes, L. (Ö), Becker, I. (1994). The complete DNA sequence of chromosome XI of Saccharomyces cerevisiae Nature 369 :371-378

Bossier, P., Fernandes, L., Rocha, D., Rodrigues-Pousada, C. (1993). Overexpression of YAP2, coding for a new yAP protein, and YAP1 in Saccharomyces cerevisiae alleviates growth inhibition caused by 1,10-phenantroline J. Biol. Chem 268 :23640-23645

Bossier, P., Fernandes, L., Rodrigues-Pousada, C. (1992). Yeast genes overcoming growth arrest induced by 1,10-phenantroline Cell Biology 69 :7-11