CBMA

The research group activities span from molecules to cells and whole organisms. The group possesses a solid know-how on physiology, biochemistry and molecular biology, with applications in the fields of biotechnology and biomedicine.

The team exploits cutting-edge technologies established for the budding yeast Saccharomyces cerevisiae to investigate and characterize molecular and cellular targets of toxic compounds, and integrates this information to uncover novel functional networks and pathways in key biological processes, which are evolutionary conserved. Thus, we could also translate our findings to different biological models, contributing to the understanding of mechanisms underlying the functioning of higher eukaryotic cells.

Main achievements

The RG contributed significantly to highlight several signalling and regulatory pathways of cellular processes triggered by biotic and abiotic stressors, with special focus on plasma membrane (PM) transporters, programmed cell death (PCD) and host-fungus adaptive responses.

Understanding the complex PCD regulatory network has been a key research topic of the RG. We identified common features between mitochondria death pathways in yeast and mammals, contributing to the recognition that PCD is highly conserved (1). We also heterologously (co-)expressed the pro-apoptotic mammalian Bax to elucidate the regulatory PCD network of higher eukaryotes (2). More recently, we also used the yeast model to uncover ammonium as an ageing determinant (3,4, 5) and cell death induced by cisplatin (6) as well as to identify the role of ceramide metabolism in apoptosis through the modulation of mitochondrial permeabilization (7,8). Our study of the cytotoxic effect of C2-phytoceramide revealed that this synthetic sphingolipid perturbs lipid rafts and cell integrity in S. cerevisiae in a sterol-dependent manner (9). In another work we uncovered that the protective role of yeast cathepsin D in acetic acid-induced apoptosis (10) and found it depends on ANT (Aac2p) but not on the voltage-dependent channel (Por1p) (11). Furthermore, we proposed that the vacuolar release of the cathepsin D also regulates PCD in colorectal carcinoma (CRC), and exploited this model in studies of signaling pathways involved in carcinogenesis and resistance to therapy (12,13). Aiming to elucidate the mechanisms underlying lysosomal induced cell death we also optimise several flow cytometric protocols to determine dysfunction of the yeast like lysosome in purified fractions of this organelle (14). With the goal to further exploit our well-established acetic acid-induced cell death model we performed a genome-wide approach and identified genes involved in the positive and negative regulation of acetic acid-induced programmed cell death in S. cerevisiae (15).

The characterization of plasma membrane (PM) transporters has been an internationally recognized key research theme of the RG in the last 25 years. PM proteins intervene on how the cell interacts with its surroundings, including sensing, adhesion, signaling and solute uptake, which allow the cell to respond to distinct environmental cues. We achieved important milestones in this field, including the characterization of PM lactate transporters in different yeast species (16) in breast cancer cell lines (17), and bacteria (18). PM proteins are tightly regulated and are rapidly targeted for vacuolar degradation, which depends on phosphorylation, ubiquitylation and endocytosis (19). Recently, we showed the activation of arrestin-related proteins is involved in the mediation of intracellular signaling (20).

The ability of the yeast C. albicans to escape/adjust to host defences depends on its dynamic interactions with the host, in which virulence factors like proteolytic enzymes or cell wall determinants were claimed to play a signi?cant role. We showed that secreted aspartyl proteinases have a limited role in the murine hematogenously disseminated infection but are important for other types of infections (21). Furthermore, we revealed for the first time that continuous interaction with the host leads to decreased strain virulence in the murine model of disseminated candidiasis (22), which is modulated by the cell wall integrity signaling pathway. Our recent results suggest a complex regulation, by RLM1, of cell wall determinants and virulence factors, providing the possibility to exploit these pathways and their interplay to unravel fungal adaptive processes.

Overall, we characterized PM proteins, PCD regulators and virulence factors that are attractive molecular targets to develop novel therapeutics for Human pathologies associated with cell death dysfunctions and infectious diseases, as well as to improve yeast bio-based industrial processes. The knowledge generated will contribute to uncover signaling/regulatory pathways of cellular events triggered by biotic and abiotic stressors.

Key publications (last 5 years)

1. Pereira, C., Silva, R.D., Saraiva, L., Johansson, B., Sousa, M.J. and Côrte-Real, M. (2008) Mitochondria dependent apoptosis in yeast. Biochim Biophys Acta 1783(7), 1286-302.

2 - Silva RD, Manon S, Gonçalves J, Saraiva L, Côrte-Real M. (2011). The importance of humanized yeast to better understand the role of bcl-2 family in apoptosis: finding of novel therapeutic opportunities. Current Pharmaceutical Design, 17: 246-55. DOI: 10.2174/138161211795049651

3 - Santos J, Leão C, Sousa MJ. (2012). Growth culture conditions and nutrient signaling modulating yeast chronological longevity. Oxid Med Cell Longev, 2012: 680304. DOI: 10.1155/2012/680304

4 - Santos, J., Sousa, M.J. and Leão, C. (2012) Ammonium is toxic for aging yeast cells, inducing death and shortening of the chronological lifespan. PLOS ONE 7(5):e37090. doi: 10.1371/journal.pone.0037090.

5 - Santos, J., Leão, C. and Sousa, M.J. (2013) Ammonium-Dependent Shortening of CLS in Yeast Cells Starved for Essential Amino Acids Is Determined by the Specific Amino Acid Deprived, through Different Signaling Pathways. Oxidative Medicine and Cellular Longevity, vol. 2013, Article ID 161986

6 - Cunha D., Cunha R., Côrte-Real M. and Chaves S. R. (2013). Cisplatin-induced cell death in Saccharomyces cerevisiae is programmed and rescued by proteasome inhibition. DNA Repair (Amst). 2013 Jun 1;12(6):444-9.

7 - Rego A, Costa M, Chaves SR, Matmati N, Pereira H, Sousa MJ, Moradas-Ferreira P, Hannun YA, Costa V, Côrte-Real M. (2012). Modulation of mitochondrial outer membrane permeabilization and apoptosis by ceramide metabolism. PLoS ONE, 7: e48571.   DOI: 10.1371/journal.pone.0048571

8 - Rego A, Trindade D, Chaves S, Manon S, Costa V, Sousa MJ, Côrte-Real M (2013) The yeast model system as a tool towards the understanding of apoptosis regulation by sphingolipids, FEMS Yeast Research in production DOI: 10.1111/1567-1364.12096.

9 - Pacheco A, Azevedo F, Rego A, Santos J, Chaves SR, Côrte-Real M and Sousa MJ C2-phytoceramide perturbs lipid rafts and cell integrity in Saccharomyces cerevisiae in a sterol-dependent manner (2013) PLoS ONE in production (IF 2012: 3.73).

10 - Pereira, C., Chaves, S., Alves, S., Salin, B., Camougrand, N., Manon, S., Sousa, M.J. and Côrte-Real, M. (2010) Mitochondrial degradation in acetic acid-induced yeast apoptosis: the role of Pep4 and the ADP/ATP carrier. Mol Microbiol 76(6), 1398-410.

11 - Pereira H., Azevedo F., Rego A., Sousa M.J., Chaves, S.R. and Côrte-Real M. (2013) The protective role of yeast cathepsin D in acetic acid-induced apoptosis depends on ANT (Aac2p) but not on the voltage-dependent channel (Por1p) FEBS Letters 587, 200–205.

12 - Marques C, Oliveira C.S. F., Alves S., Chaves S.R., Coutinho OP, Côrte-Real M and Preto A. Acetate-induced apoptosis in colorectal carcinoma cells involves lysosomal membrane permeabilization and cathepsin D release. Cell Death Dis. 2013 Feb 21;4.

13 - Preto A, Figueiredo J, Velho S, Ribeiro AS, Soares P, Oliveira C, Seruca R. (2008). BRAF provides proliferation and survival signals in MSI colorectal carcinoma cells displaying BRAFV600E but not KRAS mutations. The Journal of Pathology, 214: 320-327. DOI: 10.1002/path.2295

14 - Rodrigues J, Silva RD, Noronha H, Pedras A, Gerós H, Côrte-Real M (2013) Flow cytometry as a novel tool for structural and functional characterization of isolated yeast vacuoles. Microbiology 159 (Pt 5):848-56.

15 - Sousa M, Duarte AM, Fernandes TR, Chaves SR, Pacheco A, Leão C, Côrte-Real M, Sousa MJ. Genome-wide identification of genes involved in the positive and negative regulation of acetic acid-induced programmed cell death in Saccharomyces cerevisiae. BMC Genomics. 2013 Nov 28;14:838. doi: 10.1186/1471-2164-14-838

16 - Casal M, Paiva S, Queirós O, Soares-Silva I. (2008). Transport of carboxylic acids in yeasts. FEMS Microbiology Reviews, 32: 974-994. DOI: 10.1111/j.1574-6976.2008.00128.x  

17 - Queirós O, Preto A, Pacheco A, Pinheiro C, Azevedo-Silva J, Moreira R, Pedro M, Ko YH, Pedersen PL, Baltazar F, Casal M. (2012). Butyrate activates the monocarboxylate transporter MCT4 expression in breast cancer cells and enhances the antitumor activity of 3-bromopyruvate. J.Bioenerg. Biomembr, 44: 141-153. DOI: 10.1007/s10863-012-9418-3

18 – Sá-Pessoa J, Paiva S, Ribas D, Silva IJ, Viegas SC, Arraiano CM, Casal M. (2013) SATP (YaaH), a succinate-acetate transporter protein in Escherichia coli. Biochem J. 2013 Sep 15;454(3):585-95. doi: 10.1042/BJ20130412

19 - Paiva, S, Vieira, N, Nondier, I, Haguenauer-Tsapis, R, Casal, M, Urban-Grimal, D. (2009). Glucose-induces ubiquitylation and endocytosis of the yeast Jen1 transporter: role of ubiquitin-K63 chains. Journal of Biological Chemistry, 284: 19228-19236. DOI:  10.1074/jbc.M109.008318

20 - Becuwe M, Vieira N, Lara D, Gomes-Rezende J, Soares-Cunha C, Haguenauer-Tsapis R, Vincent O, Casal M, Paiva S, Léon S. (2012). A molecular switch on an arrestin-like protein relays glucose signaling to transporter endocytosis. Journal of Cell Biology, 196: 247-55. DOI: 10.1083/jcb.201109113  

21 – Sabino R, Veríssimo C, Brandao J, Alves C, Parada H, Rosado L, Paixao E, Videira Z, Tendeiro T, Sampaio P, Pais C. (2010). Epidemiology of candidemia in oncology patients: a 6-year survey in a Portuguese central hospital. Medical Mycology, 48: 346-354. DOI: 10.3109/13693780903161216

22 - Sampaio P, Santos M, Correia A, Amaral FE, Chavez-Galarza J, Costa-de-Oliveira S, Castro AG, Pedrosa J, Pais C. (2010). Virulence Attenuation of Candida albicans Genetic Variants Isolated from a Patient with a Recurrent Bloodstream Infection. Plos One, 5: e10155. DOI: 10.1371/journal.pone.0010155