High rate of A(H1N1)pdm09 infections among rural Thai villagers, 2009-2010

10.1371/journal.pone.0106751PLoS ONE99e10675

Prospective Study of Avian Influenza Virus Infections among Rural Thai Villagers

10.1371/journal.pone.0072196PLoS ONE88e7219

Loss of ATRX, Genome Instability, and an Altered DNA Damage Response Are Hallmarks of the Alternative Lengthening of Telomeres Pathway

<div><p>The Alternative Lengthening of Telomeres (ALT) pathway is a telomerase-independent pathway for telomere maintenance that is active in a significant subset of human cancers and in vitro immortalized cell lines. ALT is thought to involve templated extension of telomeres through homologous recombination, but the genetic or epigenetic changes that unleash ALT are not known. Recently, mutations in the ATRX/DAXX chromatin remodeling complex and histone H3.3 were found to correlate with features of ALT in pancreatic neuroendocrine cancers, pediatric glioblastomas, and other tumors of the central nervous system, suggesting that these mutations might contribute to the activation of the ALT pathway in these cancers. We have taken a comprehensive approach to deciphering ALT by applying genomic, molecular biological, and cell biological approaches to a panel of 22 ALT cell lines, including cell lines derived in vitro. Here we show that loss of ATRX protein and mutations in the <em>ATRX</em> gene are hallmarks of ALT–immortalized cell lines. In addition, ALT is associated with extensive genome rearrangements, marked micronucleation, defects in the G2/M checkpoint, and altered double-strand break (DSB) repair. These attributes will facilitate the diagnosis and treatment of ALT positive human cancers.</p> </div

Description of ALT lines used in this study and summary of results.

(a)<p>CF, cystic fibrosis; HPV, human papillovirus 16 E6 and E7; LF, Li-Fraumeni syndrome; LN, Lesch-Nyhan syndrome; OS, osteosarcoma; SV40 ER, simian virus 40 early region.</p>(b)<p>Values are averages of triplicate assays (±SD) with JFCF-6/T.1D set to 100 in each assay and the other values expressed relative to this standard. BJ and HeLa values are below 2.</p>(c)<p>Bold: aberrant ATRX or DAXX protein in western and/or IF. ATRX western results are from three independent immunoblots. Examples of abnormal IF are given in <a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1002772#pgen-1002772-g001" target="_blank">Figure 1b</a>.</p>(d)<p>Bold: deletions in ATRX. See <a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1002772#pgen.1002772.s008" target="_blank">Table S1</a> for details.</p>(e)<p>Values represent mean % cells with micronuclei and SDs from three independent experiments. See <a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1002772#pgen-1002772-g004" target="_blank">Figure 4</a>. Values for HeLa and BJ/SV40 are <8%. Bold: >10% of cells with micronuclei.</p>(f)<p>Bold: abnormal G2/M checkpoint initiation (<70% reduction in mitotic index 1 hr after 10 Gy IR). See <a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1002772#pgen-1002772-g005" target="_blank">Figure 5A</a>. Italic: uninterpretable due to low mitotic index. hTERT-RPE (positive control) 89% and ATM-/- GM5849 46%.</p>(g)<p>Bold: abnormal G2/M checkpoint maintenance (<90% reduction in mitotic index 16 hr after 4.5 Gy IR). See <a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1002772#pgen-1002772-g005" target="_blank">Figure 5b</a>. Italic: low mitotic index at 16 hr in noc. Values for HeLa and BJ/SV40 were 98±2% and 96±3%, respectively.</p>(h)<p>Bold: slow DSB repair kinetics. See <a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1002772#pgen-1002772-g006" target="_blank">Figure 6A</a> and Figure S8. After 0.5 Gy IR, cells with >10 53BP1 foci were scored at 1 and 24 hr and without IR. Values represent (% at 24 hr)-(% no IR)/(% at 1 hr)-(% no IR). nt: not tested.</p>(i)<p>Bold: abnormal residual DNA fragmentation 24 hr post IR. See <a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1002772#pgen-1002772-g006" target="_blank">Figure 6B</a>. nt: not tested.</p>(j)<p>Bold: greater than 60% of cells containing >10 spontaneous 53BP1 foci.</p>(k)<p>A. Englezou, P. Bonnefin, R. Reddel, unpublished data.</p>(l)<p>J. Plowman, L. Huschtscha, R. Reddel, unpublished data.</p

FAN1 mutations cause karyomegalic interstitial nephritis, linking chronic kidney failure to defective DNA damage repair

Chronic kidney disease (CKD) represents a major health burden. Its central feature of renal fibrosis is not well understood. By exome sequencing, we identified mutations in FAN1 as a cause of karyomegalic interstitial nephritis (KIN), a disorder that serves as a model for renal fibrosis. Renal histology in KIN is indistinguishable from that of nephronophthisis, except for the presence of karyomegaly. The FAN1 protein has nuclease activity and acts in DNA interstrand cross-link (ICL) repair within the Fanconi anemia DNA damage response (DDR) pathway. We show that cells from individuals with FAN1 mutations have sensitivity to the ICL-inducing agent mitomycin C but do not exhibit chromosome breakage or cell cycle arrest after diepoxybutane treatment, unlike cells from individuals with Fanconi anemia. We complemented ICL sensitivity with wild-type FAN1 but not with cDNA having mutations found in individuals with KIN. Depletion of fan1 in zebrafish caused increased DDR, apoptosis and kidney cysts. Our findings implicate susceptibility to environmental genotoxins and inadequate DNA repair as novel mechanisms contributing to renal fibrosis and CKD

Abnormal karyotypes in ALT lines.

<p>Representative metaphases from 5 cell lines showing subtetraploid karyotype with high number of rearranged chromosomes (up to 60%). Structural rearrangements are labeled as following: del(Z)- deletion of chromosome Z; der(Z) - multiple aberrations within single chromosome Z; chromosome, denoted with two or more numbers indicate rearrangement involving two or more partners.</p