Worldwide, several million workers are employed in the various chromium (Cr) industries. These workers may suffer from a variety of adverse health effects produced by dusts, mists and fumes containing Cr in the hexavalent oxidation state, Cr(VI). Of major importance, occupational exposure to Cr(VI) compounds has been firmly associated with the development of lung cancer. Counterintuitively, Cr(VI) is mostly unreactive towards most biomolecules, including nucleic acids. However, its intracellular reduction produces several species that react extensively with biomolecules. The diversity and chemical versatility of these species add great complexity to the study of the molecular mechanisms underlying Cr(VI) toxicity and carcinogenicity. As a consequence, these mechanisms are still poorly understood, in spite of intensive research efforts. Here, we discuss the impact of Cr(VI) on the stress response—an intricate cellular system against proteotoxic stress which is increasingly viewed as playing a critical role in carcinogenesis. This discussion is preceded by information regarding applications, chemical properties and adverse health effects of Cr(VI). A summary of our current understanding of cancer initiation, promotion and progression is also provided, followed by a brief description of the stress response and its links to cancer and by an overview of potential molecular mechanisms of Cr(VI) carcinogenicity.

DNA oxidation by ten-eleven translocation (TET) family enzymes is essential for epigenetic reprogramming. The conversion of 5-methylcytosine (5mC) into 5-hydroxymethylcytosine (5hmC) initiates developmental and cell-type-specific transcriptional programs through mechanisms that include changes in the chromatin structure. Here, we show that the presence of 5hmC in the transcribed gene promotes the annealing of the nascent RNA to the template DNA strand, leading to the formation of an R-loop. Depletion of TET enzymes reduced global R-loops in the absence of gene expression changes, whereas CRISPR-mediated tethering of TET to an active gene promoted the formation of R-loops. The genome-wide distribution of 5hmC and R-loops shows a positive correlation in mouse and human stem cells and overlap in half of all active genes. Moreover, R-loop resolution leads to differential expression of a subset of genes that are involved in crucial events during stem cell proliferation. Altogether, our data reveal that epigenetic reprogramming via TET activity promotes co-transcriptional R-loop formation, disclosing new mechanisms of gene expression regulation.

RNA-protein interactions drive key cellular pathways such as protein translation, nuclear organization and genome stability maintenance. The human telomeric protein TRF2 binds to the long noncoding RNA TERRA through independent domains, including its N-terminal B domain. We previously demonstrated that TRF2 B domain binding to TERRA supports invasion of TERRA into telomeric double stranded DNA, leading to the formation of telomeric RNA:DNA hybrids. The other telomeric protein TRF1, which also binds to TERRA, suppresses this TRF2-associated activity by preventing TERRA-B domain interactions. Herein, we show that the binding of both TRF1 and TRF2 to TERRA depends on the ability of the latter to form G-quadruplex structures. Moreover, a cluster of arginines within the B domain is largely responsible for its binding to TERRA. On the other side, a patch of glutamates within the N-terminal A domain of TRF1 mainly accounts for the inhibition of TERRA-B domain complex formation. Finally, mouse TRF2 B domain binds to TERRA, similarly to its human counterpart, while mouse TRF1 A domain lacks the inhibitory activity. Our data shed further light on the complex crosstalk between telomeric proteins and RNAs and suggest a lack of functional conservation in mouse.

Cancer cells with an activated Alternative Lengthening of Telomeres (ALT) mechanism elongate telomeres via homology-directed repair. Sustained telomeric replication stress is an essential trigger of ALT activity; however, it can lead to cell death if not properly restricted. By analyzing publicly available data from genome-wide CRISPR KO screenings, we have identified the multifunctional protein PC4 as a novel factor essential for ALT cell viability. Depletion of PC4 results in rapid ALT cell death, while telomerase-positive cells show minimal effects. PC4 depletion induces replication stress and telomere fragility primarily in ALT cells, and increases ALT activity. PC4 binds to telomeric DNA in cells, and its binding can be enhanced by telomeric replication stress. Finally, a mutant PC4 with partly impaired single stranded DNA binding activity is capable to localize to telomeres and suppress ALT activity and telomeric replication stress. We propose that PC4 supports ALT cell viability, at least partly, by averting telomere dysfunction. Further studies of PC4 interactions at ALT telomeres may hold promise for innovative therapies to eradicate ALT cancers.

In the 1920s, Otto Warburg, one of the leading biochemists of the 20th century, uncovered a striking phenotype of cancer cells: their increased dependence on lactic acid fermentation for energy production compared to that of the normal cells from which they derived. Warburg viewed this metabolic particularity of cancer cells, which came to be known as the Warburg effect, as a driving force in carcinogenesis. This perception suggested a novel path for cancer therapy, a strategy that Warburg himself proposed and defended with passion to his death. However, for many decades, both his metabolic theory of cancer and suggested therapeutic approach were essentially ignored by cancer researchers, who were mostly focused on the genetic basis of the disease and on the intricacies of the pathways known to promote cellular proliferation, differentiation and death. Still, thanks to the combined efforts of those who chose to pursue Warburg’s line of research, experimental evidence supporting and extending Warburg´s findings on the metabolism of cancer cells accumulated. In the 1980s, 18F-deoxyglucose positron emission tomography (18FDG–PET) was implemented in the clinic. This metabolic imaging technique, which is based on the avidity of cancer cells for glucose, represents, to this day, the only successful exploitation of the Warburg effect for medical purposes. The wide success of 18FDG–PET in the diagnosis and staging of tumors is among the factors most responsible for renewing interest in the central carbon metabolism of cancer cells. This renewed interest was further boosted by the discovery of multiple links between central carbon metabolism and cellular proliferation, differentiation and death and culminated in the recent classification, by Weinberg and Hanahan, of tumor metabolism as an emerging cancer hallmark. Tremendous research effort is now being devoted into a more detailed and comprehensive elucidation of the metabolic rewiring that accompanies neoplastic transformation and, unsurprisingly, targeting the metabolic peculiarities of tumors has become a hot topic in drug discovery. This chapter summarizes past and current efforts at targeting the Warburg effect for selective cancer therapies.