75 research outputs found
Stem Cell-Specific Mechanisms Ensure Genomic Fidelity within HSCs and upon Aging of HSCs
SummaryWhether aged hematopoietic stem and progenitor cells (HSPCs) have impaired DNA damage repair is controversial. Using a combination of DNA mutation indicator assays, we observe a 2- to 3-fold increase in the number of DNA mutations in the hematopoietic system upon aging. Young and aged hematopoietic stem cells (HSCs) and hematopoietic progenitor cells (HPCs) do not show an increase in mutation upon irradiation-induced DNA damage repair, and young and aged HSPCs respond very similarly to DNA damage with respect to cell-cycle checkpoint activation and apoptosis. Both young and aged HSPCs show impaired activation of the DNA-damage-induced G1-S checkpoint. Induction of chronic DNA double-strand breaks by zinc-finger nucleases suggests that HSPCs undergo apoptosis rather than faulty repair. These data reveal a protective mechanism in both the young and aged hematopoietic system against accumulation of mutations in response to DNA damage
The human DEK oncogene regulates DNA damage response signaling and repair
The human DEK gene is frequently overexpressed and sometimes amplified in human cancer. Consistent with oncogenic functions, Dek knockout mice are partially resistant to chemically induced papilloma formation. Additionally, DEK knockdown in vitro sensitizes cancer cells to DNA damaging agents and induces cell death via p53-dependent and -independent mechanisms. Here we report that DEK is important for DNA double-strand break repair. DEK depletion in human cancer cell lines and xenografts was sufficient to induce a DNA damage response as assessed by detection of γH2AX and FANCD2. Phosphorylation of H2AX was accompanied by contrasting activation and suppression, respectively, of the ATM and DNA-PK pathways. Similar DNA damage responses were observed in primary Dek knockout mouse embryonic fibroblasts (MEFs), along with increased levels of DNA damage and exaggerated induction of senescence in response to genotoxic stress. Importantly, Dek knockout MEFs exhibited distinct defects in non-homologous end joining (NHEJ) when compared to their wild-type counterparts. Taken together, the data demonstrate new molecular links between DEK and DNA damage response signaling pathways, and suggest that DEK contributes to DNA repair
Dipoid-Specific Genome Stability Genes of S. cerevisiae: Genomic Screen Reveals Haploidization as an Escape from Persisting DNA Rearrangement Stress
Maintaining a stable genome is one of the most important tasks of every living cell and the mechanisms ensuring it are similar in all of them. The events leading to changes in DNA sequence (mutations) in diploid cells occur one to two orders of magnitude more frequently than in haploid cells. The majority of those events lead to loss of heterozygosity at the mutagenesis marker, thus diploid-specific genome stability mechanisms can be anticipated. In a new global screen for spontaneous loss of function at heterozygous forward mutagenesis marker locus, employing three different mutagenesis markers, we selected genes whose deletion causes genetic instability in diploid Saccharomyces cerevisiae cells. We have found numerous genes connected with DNA replication and repair, remodeling of chromatin, cell cycle control, stress response, and in particular the structural maintenance of chromosome complexes. We have also identified 59 uncharacterized or dubious ORFs, which show the genome instability phenotype when deleted. For one of the strongest mutators revealed in our screen, ctf18Δ/ctf18Δ the genome instability manifests as a tendency to lose the whole set of chromosomes. We postulate that this phenomenon might diminish the devastating effects of DNA rearrangements, thereby increasing the cell's chances of surviving stressful conditions. We believe that numerous new genes implicated in genome maintenance, together with newly discovered phenomenon of ploidy reduction, will help revealing novel molecular processes involved in the genome stability of diploid cells. They also provide the clues in the quest for new therapeutic targets to cure human genome instability-related diseases
Cell cycle specific fluctuations in adenosine 3':5'-cyclic monophosphate and polyamines of Chinese hamster cells.
Nuclear receptor LRH-1/NR5A2 is required and targetable for liver endoplasmic reticulum stress resolution
Chronic endoplasmic reticulum (ER) stress results in toxicity that contributes to multiple human disorders. We report a stress resolution pathway initiated by the nuclear receptor LRH-1 that is independent of known unfolded protein response (UPR) pathways. Like mice lacking primary UPR components, hepatic Lrh-1-null mice cannot resolve ER stress, despite a functional UPR. In response to ER stress, LRH-1 induces expression of the kinase Plk3, which phosphorylates and activates the transcription factor ATF2. Plk3-null mice also cannot resolve ER stress, and restoring Plk3 expression in Lrh-1-null cells rescues ER stress resolution. Reduced or heightened ATF2 activity also sensitizes or desensitizes cells to ER stress, respectively. LRH-1 agonist treatment increases ER stress resistance and decreases cell death. We conclude that LRH-1 initiates a novel pathway of ER stress resolution that is independent of the UPR, yet equivalently required. Targeting LRH-1 may be beneficial in human disorders associated with chronic ER stress
Identification of DNA sequences required for mouse APRT gene expression.
The mouse aprt promoter contains four GC boxes, which bind transcription factor Spl in vitro, and lacks both TATA and CCAAT boxes. Removal of the two most distal GC boxes of this promoter had little effect on APRT enzyme levels produced in a transient expression assay. Deletion of the distal three GC boxes resulted in a 50% reduction, and deletion of all GC boxes resulted in essentially complete loss of APRT activity. There are two predominant transcription start sites which are located within the region containing the GC boxes. The promoter behaved as a relatively strong promoter when compared to the RSV LTR promoter in a transient CAT assay, and operated in one orientation only. No upstream anti-sense transcripts were detected in either mouse CAK or liver cells, confirming that the mouse aprt promoter, unlike some other GC-rich promoters appears not to support bidirectional transcription
Comparative anatomy of the human APRT gene and enzyme: nucleotide sequence divergence and conservation of a nonrandom CpG dinucleotide arrangement.
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