Acetic acid stress in budding yeast: From molecular mechanisms to applications

Acetic acid stress represents a frequent challenge to counteract for yeast cells under several environmental conditions and industrial bioprocesses. The molecular mechanisms underlying its response have been mostly elucidated in the budding yeast Saccharomyces cerevisiae, where acetic acid can be either a physiological substrate or a stressor. This review will focus on acetic acid stress and its response in the context of cellular transport, pH homeostasis, metabolism and stress-signalling pathways. This information has been integrated with the results obtained by multi-omics, synthetic biology and metabolic engineering approaches aimed to identify major cellular players involved in acetic acid tolerance. In the production of biofuels and renewable chemicals from lignocellulosic biomass, the improvement of acetic acid tolerance is a key factor. In this view, how this knowledge could be used to contribute to the development and competitiveness of yeast cell factories for sustainable applications will be also discussed

Yeast as a Tool to Study Signaling Pathways in Mitochondrial Stress Response and Cytoprotection

Cell homeostasis results from the balance between cell capability to adapt or succumb to environmental stress. Mitochondria, in addition to supplying cellular energy, are involved in a range of processes deciding about cellular life or death. The crucial role of mitochondria in cell death is well recognized. Mitochondrial dysfunction has been associated with the death process and the onset of numerous diseases. Yet, mitochondrial involvement in cellular adaptation to stress is still largely unexplored. Strong interest exists in pharmacological manipulation of mitochondrial metabolism and signaling. The yeast Saccharomyces cerevisiae has proven a valuable model organism in which several intracellular processes have been characterized in great detail, including the retrograde response to mitochondrial dysfunction and, more recently, programmed cell death. In this paper we review experimental evidences of mitochondrial involvement in cytoprotection and propose yeast as a model system to investigate the role of mitochondria in the cross-talk between prosurvival and prodeath pathways

The Transcription Factors ADR1 or CAT8 are Required for RTG Pathway Activation and Evasion from Yeast Acetic Acid-Induced Programmed Cell Death in Raffinose

Yeast Saccharomyces cerevisiae grown on glucose undergoes programmed cell death (PCD) induced by acetic acid (AA-PCD), but evades PCD when grown in raffinose. This is due to concomitant relief of carbon catabolite repression (CCR) and activation of mitochondrial retrograde signaling, a mitochondria-to-nucleus communication pathway causing up-regulation of various nuclear target genes, such as CIT2, encoding peroxisomal citrate synthase, dependent on the positive regulator RTG2 in response to mitochondrial dysfunction. CCR down-regulates genes mainly involved in mitochondrial respiratory metabolism. In this work, we investigated the relationships between the RTG and CCR pathways in the modulation of AA-PCD sensitivity under glucose repression or de-repression conditions. Yeast single and double mutants lacking RTG2 and/or certain factors regulating carbon source utilization, including MIG1, HXK2, ADR1, CAT8, and HAP4, have been analyzed for their survival and CIT2 expression after acetic acid treatment. ADR1 and CAT8 were identified as positive regulators of RTG-dependent gene transcription. ADR1 and CAT8 interact with RTG2 and with each other in inducing cell resistance to AA-PCD in raffinose and controlling the nature of cell death. In the absence of ADR1 and CAT8, AA-PCD evasion is acquired through activation of an alternative factor/pathway repressed by RTG2, suggesting that RTG2 may play a function in promoting necrotic cell death in repressing conditions when RTG pathway is inactive. Moreover, our data show that simultaneous mitochondrial retrograde pathway activation and SNF1-dependent relief of CCR have a key role in central carbon metabolism reprogramming which modulates the yeast acetic acid-stress response

The transcription factors ADR1 or CAT8 are required for RTG pathway activation and evasion from yeast acetic acid-induced programmed cell death in raffinose

Yeast Saccharomyces cerevisiae grown on glucose undergoes programmed cell death (PCD) induced by acetic acid (AA-PCD), but evades PCD when grown in raffinose. This is due to concomitant relief of carbon catabolite repression (CCR) and activation of mitochondrial retrograde signaling, a mitochondria-to-nucleus communication pathway causing up-regulation of various nuclear target genes, such as CIT2, encoding peroxisomal citrate synthase, dependent on the positive regulator RTG2 in response to mitochondrial dysfunction. CCR down-regulates genes mainly involved in mitochondrial respiratory metabolism. In this work, we investigated the relationships between the RTG and CCR pathways in the modulation of AA-PCD sensitivity under glucose repression or de-repression conditions. Yeast single and double mutants lacking RTG2 and/or certain factors regulating carbon source utilization, including MIG1, HXK2, ADR1, CAT8, and HAP4, have been analyzed for their survival and CIT2 expression after acetic acid treatment. ADR1 and CAT8 were identified as positive regulators of RTG-dependent gene transcription. ADR1 and CAT8 interact with RTG2 and with each other in inducing cell resistance to AA-PCD in raffinose and controlling the nature of cell death. In the absence of ADR1 and CAT8, AA-PCD evasion is acquired through activation of an alternative factor/pathway repressed by RTG2, suggesting that RTG2 may play a function in promoting necrotic cell death in repressing conditions when RTG pathway is inactive. Moreover, our data show that simultaneous mitochondrial retrograde pathway activation and SNF1-dependent relief of CCR have a key role in central carbon metabolism reprogramming which modulates the yeast acetic acid-stress response

New perspectives from South-Y-East, not all about death. A report of the 12lnternational Meeting on Yeast Apoptosis in Bari, Italy, May 14th-18th, 2017

Over the last 14 years, the field of yeast regulated cell death (RCD) has been expanding to more and more biomedical research themes, including aging, human diseases, cell stress response, metabolism and systems biology. The 12th International Meeting on Yeast Apoptosis (IMYA12), which was held in Bari, Italy from May 14th to 18th, 2017, nicely reflected this trend. This year, more than 100 participants, among which senior and young scientists from Europe, USA, North Africa and Japan, had an intense and open exchange of achievements and ideas. This open and informal communication among researchers has been a constant hallmark of all the IMYA meetings. The global yeast death community was embraced and inspired by the lively and warm atmosphere of Bari, the capital city of Apulia, and its beautiful surroundings, with colorful landscapes, historical and artistic heritage, tastes and scents that reflect the interface between Eastern and Western culture

Guidelines and Recommendations on Yeast Cell Death Nomenclature

Elucidating the biology of yeast in its full complexity has major implications for science, medicine and industry. One of the most critical processes determining yeast life and physiology is cellular demise. However, the investigation of yeast cell death is a relatively young field, and a widely accepted set of concepts and terms is still missing. Here, we propose unified criteria for the definition of accidental, regulated, and programmed forms of cell death in yeast based on a series of morphological and biochemical criteria. Specifically, we provide consensus guidelines on the differential definition of terms including apoptosis, regulated necrosis, and autophagic cell death, as we refer to additional cell death routines that are relevant for the biology of (at least some species of) yeast. As this area of investigation advances rapidly, changes and extensions to this set of recommendations will be implemented in the years to come. Nonetheless, we strongly encourage the authors, reviewers and editors of scientific articles to adopt these collective standards in order to establish an accurate framework for yeast cell death research and, ultimately, to accelerate the progress of this vibrant field of research

Guidelines and recommendations on yeast cell death nomenclature

Elucidating the biology of yeast in its full complexity has major implications for science, medicine and industry. One of the most critical processes determining yeast life and physiology is cel-lular demise. However, the investigation of yeast cell death is a relatively young field, and a widely accepted set of concepts and terms is still missing. Here, we propose unified criteria for the defi-nition of accidental, regulated, and programmed forms of cell death in yeast based on a series of morphological and biochemical criteria. Specifically, we provide consensus guidelines on the differ-ential definition of terms including apoptosis, regulated necrosis, and autophagic cell death, as we refer to additional cell death rou-tines that are relevant for the biology of (at least some species of) yeast. As this area of investigation advances rapidly, changes and extensions to this set of recommendations will be implemented in the years to come. Nonetheless, we strongly encourage the au-thors, reviewers and editors of scientific articles to adopt these collective standards in order to establish an accurate framework for yeast cell death research and, ultimately, to accelerate the pro-gress of this vibrant field of research

Analysis of Mitochondrial Retrograde Signaling in Yeast Model Systems

: Mitochondrial retrograde signaling is a mitochondria-to-nucleus communication pathway, conserved from yeast to humans, by which dysfunctional mitochondria relay signals that lead to cell stress adaptation in physiopathological conditions via changes in nuclear gene expression. The most comprehensive picture of components and regulation of retrograde signaling has been obtained in Saccharomyces cerevisiae, where retrograde-target gene expression is regulated by RTG genes. In this chapter, we describe methods to measure mitochondrial retrograde pathway activation at the level of mRNA and protein products in yeast model systems, including cell suspensions and microcolonies. In particular, we will focus on three major procedures: mRNA levels of RTG-target genes, such as those encoding for peroxisomal citrate synthase (CIT2), aconitase, and NAD+-specific isocitrate dehydrogenase subunit 1 by real-time PCR; expression analysis of CIT2-gene protein product (Cit2p-GFP) by Western blot and fluorescence microscopy; the phosphorylation status of transcriptional factor Rtg1/3p which controls RTG-target gene transcription

6-Thioguanine and Its Analogs Promote Apoptosis of Castration-Resistant Prostate Cancer Cells in a BRCA2-Dependent Manner

Background: Mutations in the oncosuppressor gene BReast CAncer susceptibility gene 2 (BRCA2) predispose to aggressive forms of prostate cancer which show poor response to taxane-based therapy, the standard treatment for castration-resistant, aggressive prostate cancer. Herein, we addressed the question whether changes in BRCA2 expression, a potential surrogate marker for BRCA2 activity, may affect the response of castration-resistant prostate cancer cells to 6-thioguanine (6-TG), a thiopurine used in the treatment of haematological malignancies. Methods: Yeast, normal prostate cells and castration-resistant prostate cancer cells were treated with 6-TG or its analogues, in presence or absence of paclitaxel, or with olaparib, a poly-(ADP-ribose) polymerase (PARP) inhibitor currently in clinical trials for treatment of metastatic castration-resistant prostate cancer, and cell proliferation, apoptosis and androgen receptor (AR) levels were measured. Results: 6-TG inhibited cell proliferation in yeast, normal and castration-resistant prostate cancer cells but promoted apoptosis only in cancer cells. Suppression of BRCA2 expression by siRNA or shRNA increased the sensitivity to 6-TG- and olaparib-induced apoptosis but did not affect cancer cell response to taxane. Intriguingly, 6-TG reduced AR expression levels independently on BRCA2 expression. Instead, olaparib decreased AR levels only in BRCA2-knockdown prostate cancer cells. Notably, overexpression of BRCA2 resulted in resistance of castration-resistant prostate cancer cells to 6-TG-, taxane- and olaparib-based treatment but promoted sensitivity to apoptosis induced by 2-amino-6-bromopurine and 2,6-dithiopurine, two 6-TG analogues. Conclusions: Our results provide a pre-clinical rationale for the use of 6-TG in the treatment of BRCA2-deficient castration-resistant prostate cancers, and of certain 6-TG analogues for treatment of BRCA2-proficient prostate cancers