Trabectedin (Yondelis) is a potent antitumor drug that has the unique characteristic of killing cells by poisoning the DNA nucleotide excision repair (NER) machinery. The basis for the NER-dependent toxicity has not yet been elucidated but it has been proposed as the major determinant for the drug's cytotoxicity. To study the in vivo mode of action of trabectedin and to explore the role of NER in its cytotoxicity, we used the fission yeast Schizosaccharomyces pombe as a model system. Treatment of S. pombe wild-type cells with trabectedin led to cell cycle delay and activation of the DNA damage checkpoint, indicating that the drug causes DNA damage in vivo. DNA damage induced by the drug is mostly caused by the NER protein, Rad13 (the fission yeast orthologue to human XPG), and is mainly repaired by homologous recombination. By constructing different rad13 mutants, we show that the DNA damage induced by trabectedin depends on a 46–amino acid region of Rad13 that is homologous to a DNA-binding region of human nuclease FEN-1. More specifically, an arginine residue in Rad13 (Arg961), conserved in FEN1 (Arg314), was found to be crucial for the drug's cytotoxicity. These results lead us to propose a model for the action of trabectedin in eukaryotic cells in which the formation of a Rad13/DNA-trabectedin ternary complex, stabilized by Arg961, results in cell death. (Cancer Res 2006; 66(16): 8155-62)

Transcription is a major obstacle for replication fork (RF) progression and a cause of genome instability. Part of this instability is mediated by cotranscriptional R loops, which are believed to increase by suboptimal assembly of the nascent messenger ribonucleoprotein particle (mRNP). However, no clear evidence exists that heterogeneous nuclear RNPs (hnRNPs), the basic mRNP components, prevent R-loop stabilization. Here we show that yeast Npl3, the most abundant RNA-binding hnRNP, prevents R-loop-mediated genome instability. npl3Δ cells show transcription-dependent and R-loop-dependent hyperrecombination and genome-wide replication obstacles as determined by accumulation of the Rrm3 helicase. Such obstacles preferentially occur at long and highly expressed genes, to which Npl3 is preferentially bound in wild-type cells, and are reduced by RNase H1 overexpression. The resulting replication stress confers hypersensitivity to double-strand break-inducing agents. Therefore, our work demonstrates that mRNP factors are critical for genome integrity and opens the option of using them as therapeutic targets in anti-cancer treatment.

The p53 pathway is inactivated in the majority of human cancers. Although this perturbation frequently occurs through the mutation or deletion of p53 itself, there are other mechanisms that can attenuate the pathway and contribute to tumorigenesis. For example, overexpression of important p53 negative regulators, such as murine double minute 2 (MDM2) or murine double minute 4 (MDM4), epigenetic deregulation, or even alterations in TP53 mRNA splicing. In this work, we will review the different mechanisms of p53 pathway inhibition in cancer with special focus on multiple myeloma (MM), the second most common hematological malignancy, with low incidence of p53 mutations/deletions but growing evidence of indirect p53 pathway deregulation. Translational implications for MM and cancer prognosis and treatment are also reviewed.

The UDP-glucose:glycoprotein glucosyltransferase (UGGT) is an endoplasmic reticulum sensor for quality control of glycoprotein folding.Saccharomyces cerevisiaeis the only eukaryotic organism so far described lacking UGGT-mediated transient reglucosylation of N-linked oligosaccharides. The only gene inS. cerevisiaewith similarity to those encoding UGGTs isKRE5. S. cerevisiae KRE5deletion strains show severely reduced levels of cell wall β-1,6-glucan polymer, aberrant morphology, and extremely compromised growth or lethality, depending on the strain background. Deletion of both alleles of theCandida albicans KRE5gene gives rise to viable cells that are larger than those of the wild type (WT), tend to aggregate, have enlarged vacuoles, and show major cell wall defects.C. albicans kre5/kre5mutants have significantly reduced levels of β-1,6-glucan and more chitin and β-1,3-glucan and less mannoprotein than the WT. The remaining β-1,6-glucan, about 20% of WT levels, exhibits a β-1,6-endoglucanase digestion pattern, including a branch point-to-linear stretch ratio identical to that of WT strains, suggesting that Kre5p is not a β-1,6-glucan synthase.C. albicans KRE5is a functional homologue ofS. cerevisiae KRE5; it partially complements both the growth defect and reduced cell wall β-1,6-glucan content ofS. cerevisiae kre5viable mutants.C. albicans kre5/kre5homozygous mutant strains are unable to form hyphae in several solid and liquid media, even in the presence of serum, a potent inducer of the dimorphic transition. Surprisingly the mutants do form hyphae in the presence ofN-acetylglucosamine. Finally,C. albicans KRE5homozygous mutant strains exhibit a 50% reduction in adhesion to human epithelial cells and are completely avirulent in a mouse model of systemic infection.

Ethanol consumption triggers oxidative stress by generating reactive oxygen species (ROS) through its metabolites. This process leads to steatosis and liver inflammation, which are critical for the development of alcoholic liver disease (ALD). Autophagy is a regulated dynamic process that sequesters damaged and excess cytoplasmic organelles for lysosomal degradation and may counteract the harmful effects of ROS-induced oxidative stress. These effects include hepatotoxicity, mitochondrial damage, steatosis, endoplasmic reticulum stress, inflammation, and iron overload. In liver diseases, particularly ALD, macroautophagy has been implicated as a protective mechanism in hepatocytes, although it does not appear to play the same role in stellate cells. Beyond the liver, autophagy may also mitigate the harmful effects of alcohol on other organs, thereby providing an additional layer of protection against ALD. This protective potential is further supported by studies showing that drugs that interact with autophagy, such as rapamycin, can prevent ALD development in animal models. This systematic review presents a comprehensive analysis of the literature, focusing on the role of autophagy in oxidative stress regulation, its involvement in organ–organ crosstalk relevant to ALD, and the potential of autophagy-targeting therapeutic strategies.

Lsm1 forms part of a cytoplasmic protein complex, Lsm1‐7‐Pat1, involved in the degradation of mRNAs. Here, we show that Lsm1 has an important role in promoting genomic stability in Saccharomyces cerevisiae . Budding yeast cells lacking Lsm1 are defective in recovery from replication‐fork stalling and show DNA damage sensitivity. Here, we identify histone mRNAs as substrates of the Lsm1‐7‐Pat1 complex in yeast, and show that abnormally high amounts of histones accumulate in lsm1 Δ mutant cells. Importantly, we show that the excess of histones is responsible for the lsm1 Δ replication‐fork instability phenotype, since sensitivity of lsm1 Δ cells to drugs that stall replication forks is significantly suppressed by a reduction in histone gene dosage. Our results demonstrate that improper histone stoichiometry leads to genomic instability and highlight the importance of regulating histone mRNA decay in the tight control of histone levels in yeast.