Migratory Passerine Birds as Reservoirs of Lyme Borreliosis in Europe

Birds host vector ticks and Borrelia species and vary in effectiveness as reservoirs

Myc-dependent purine biosynthesis affects nucleolar stress and therapy response in prostate cancer

The androgen receptor is a key transcription factor contributing to the development of all stages of prostate cancer (PCa). In addition, other transcription factors have been associated with poor prognosis in PCa, amongst which c-Myc (MYC) is a well-established oncogene in many other cancers. We have previously reported that the AR promotes glycolysis and anabolic metabolism; many of these metabolic pathways are also MYC-regulated in other cancers. In this study, we report that in PCa cells de novo purine biosynthesis and the subsequent conversion to XMP is tightly regulated by MYC and independent of AR activity. We characterized two enzymes, PAICS and IMPDH2, within the pathway as PCa biomarkers in tissue samples and report increased efficacy of established anti-androgens in combination with a clinically approved IMPDH inhibitor, mycophenolic acid (MPA). Treatment with MPA led to a significant reduction in cellular guanosine triphosphate (GTP) levels accompanied by nucleolar stress and p53 stabilization. In conclusion, targeting purine biosynthesis provides an opportunity to perturb PCa metabolism and enhance tumour suppressive stress responses

Acute sensitivity of DNA replication to reduction in dNTP pools following Mec1ATR inactivation

Inactivation of Mec1, the budding yeast ATR, results in a permanent S phase arrest followed by a fatal mitotic catastrophe. The mec1 S phase arrest was proposed to stem from a defect in the Mec1-Rad53-Dun1 dependent removal of Sml1, a conserved inhibitor of ribonucleotide reductase (RNR), at the onset of S phase: According to this view, Sml1 removal and the ensuing RNR activation would promote the dNTP production necessary for genome duplication. In support for this view, dNTP levels in hypomorphic mec1 or rad53 mutants and a dun1∆ strain were shown to be reduced by as much as 46% compared to a MEC1 control strain. Notably however, nearly all analyses on a lethal mec1 allele (e.g. mec1∆ or mec1-kd [kinase dead]) have been performed in a strain background that was either deleted for SML1 or over-expressing RNR1, a requirement for maintaining viability of a mutant lacking Mec1's essential function. As a result, while it is clear that absence of Mec1 causes dNTP pool to decrease, the true extent of the reduction and whether it would be sufficient to account for the replication arrest remain elusive. Here, we addressed these questions utilizing a temperature sensitive mutant, mec1-4, which maintains its viability at permissive temperature in an otherwise wild-type background, circumventing the need to exogenously manipulate Sml1 and/or RNR activity. Results show that Mec1 inactivation leads to an S phase arrest and a ~17% reduction in dNTP pool; expression of a novel suppressor, GIS2 (glucose inhibition of gluconeogenic growth suppressor 2), rescues the arrest and partially restores the dNTP pool to ~ 93% of a control. Unexpectedly modest effects of mec1 and GIS2 on dNTP levels demonstrate that the arrest does result from a severe depletion of dNTP pool as assumed, but a heightened sensitivity to its availability</p

Acute Smc5/6 depletion reveals its primary role in rDNA replication by restraining recombination at fork pausing sites

Smc5/6, a member of the conserved SMC family of complexes, is essential for growth in most organisms. Its exact functions in a mitotic cell cycle are controversial, as chronic Smc5/6 loss-of-function alleles produce varying phenotypes. To circumvent this issue, we acutely depleted Smc5/6 in budding yeast and determined the first cell cycle consequences of Smc5/6 removal. We found a striking primary defect in replication of the ribosomal DNA (rDNA) array. Each rDNA repeat contains a programmed replication fork barrier (RFB) established by the Fob1 protein. Fob1 removal improves rDNA replication in Smc5/6 depleted cells, implicating Smc5/6 in the management of programmed fork pausing. A similar improvement is achieved by removing the DNA helicase Mph1 whose recombinogenic activity can be inhibited by Smc5/6 under DNA damage conditions. DNA 2D gel analyses further show that Smc5/6 loss increases recombination structures at RFB regions; moreover, mph1 Delta and fob1 Delta similarly reduce this accumulation. These findings point to an important mitotic role for Smc5/6 in restraining recombination events when protein barriers in rDNA stall replication forks. As rDNA maintenance influences multiple essential cellular processes, Smc5/6 likely links rDNA stability to overall mitotic growth

The mutation spectrum in genomic late replication domains shapes mammalian GC content

Genome sequence compositions and epigenetic organizations are correlated extensively across multiple length scales. Replication dynamics, in particular, is highly correlated with GC content. We combine genome-wide time of replication (ToR) data, topological domains maps and detailed functional epigenetic annotations to study the correlations between replication timing and GC content at multiple scales. We find that the decrease in genomic GC content at large scale late replicating regions can be explained by mutation bias favoring A/T nucleotide, without selection or biased gene conversion. Quantification of the free dNTP pool during the cell cycle is consistent with a mechanism involving replication-coupled mutation spectrum that favors AT nucleotides at late S-phase. We suggest that mammalian GC content composition is shaped by independent forces, globally modulating mutation bias and locally selecting on functional element. Deconvoluting these forces and analyzing them on their native scales is important for proper characterization of complex genomic correlations

Rnr1, but not Rnr3, facilitates the sustained telomerase-dependent elongation of telomeres

Ribonucleotide reductase (RNR) provides the precursors for the generation of dNTPs, which are required for DNA synthesis and repair. Here, we investigated the function of the major RNR subunits Rnr1 and Rnr3 in telomere elongation in budding yeast. We show that Rnr1 is essential for the sustained elongation of short telomeres by telomerase. In the absence of Rnr1, cells harbor very short, but functional, telomeres, which cannot become elongated by increased telomerase activity or by tethering of telomerase to telomeres. Furthermore, we demonstrate that Rnr1 function is critical to prevent an early onset of replicative senescence and premature survivor formation in telomerase-negative cells but dispensable for telomere elongation by Homology-Directed-Repair. Our results suggest that telomerase has a "basal activity" mode that is sufficient to compensate for the "end-replication-problem" and does not require the presence of Rnr1 and a different "sustained activity" mode necessary for the elongation of short telomeres, which requires an upregulation of dNTP levels and dGTP ratios specifically through Rnr1 function. By analyzing telomere length and dNTP levels in different mutants showing changes in RNR complex composition and activity we provide evidence that the Mec1ATR checkpoint protein promotes telomere elongation by increasing both dNTP levels and dGTP ratios through Rnr1 upregulation in a mechanism that cannot be replaced by its homolog Rnr3

Hydroxyurea-Mediated Cytotoxicity Without Inhibition of Ribonucleotide Reductase

SummaryIn many organisms, hydroxyurea (HU) inhibits class I ribonucleotide reductase, leading to lowered cellular pools of deoxyribonucleoside triphosphates. The reduced levels for DNA precursors is believed to cause replication fork stalling. Upon treatment of the hyperthermophilic archaeon Sulfolobus solfataricus with HU, we observe dose-dependent cell cycle arrest, accumulation of DNA double-strand breaks, stalled replication forks, and elevated levels of recombination structures. However, Sulfolobus has a HU-insensitive class II ribonucleotide reductase, and we reveal that HU treatment does not significantly impact cellular DNA precursor pools. Profiling of protein and transcript levels reveals modulation of a specific subset of replication initiation and cell division genes. Notably, the selective loss of the regulatory subunit of the primase correlates with cessation of replication initiation and stalling of replication forks. Furthermore, we find evidence for a detoxification response induced by HU treatment

Migratory Passerine Birds as Reservoirs of Lyme Borreliosis in Europe

Birds host vector ticks and Borrelia species and vary in effectiveness as reservoirs.To define the role of birds as reservoirs and disseminators of Borrelia spirochetes, we characterized tick infestation and reservoir competence of migratory passerine birds in Sweden. A total of 1,120 immature Ixodes ricinus ticks were removed from 13,260 birds and assayed by quantitative polymerase chain reaction (PCR) for Borrelia, followed by DNA sequencing for species and genotype identification. Distributions of ticks on birds were aggregated, presumably because of varying encounters with ticks along migratory routes. Lyme borreliosis spirochetes were detected in 160 (14%) ticks. Borrelia garinii was the most common species in PCR-positive samples and included genotypes associated with human infections. Infestation prevalence with infected ticks was 5 times greater among ground-foraging birds than other bird species, but the 2 groups were equally competent in transmitting Borrelia. Migratory passerine birds host epidemiologically important vector ticks and Borrelia species and vary in effectiveness as reservoirs on the basis of their feeding behavior