A role for Separase in telomere protection

Drosophila telomeres are elongated by transposition of specialized retroelements rather than telomerase activity and are assembled independently of the sequence. Fly telomeres are protected by the terminin complex that localizes and functions exclusively at telomeres and by non-terminin proteins that do not serve telomere-specific functions. We show that mutations in the Drosophila Separase encoding gene Sse lead not only to endoreduplication but also telomeric fusions (TFs), suggesting a role for Sse in telomere capping. We demonstrate that Separase binds terminin proteins and HP1, and that it is enriched at telomeres. Furthermore, we show that loss of Sse strongly reduces HP1 levels, and that HP1 overexpression in Sse mutants suppresses TFs, suggesting that TFs are caused by a HP1 diminution. Finally, we find that siRNA-induced depletion of ESPL1, the Sse human orthologue, causes telomere dysfunction and HP1 level reduction in primary fibroblasts, highlighting a conserved role of Separase in telomere protection

Deciphering the molecular machinery of stem cells: a look at the neoblast gene expression profile

Comparison of the gene-expression profiles of planarians in which all adult pluripotent stem cells (neoblasts) were eliminated and wild-type worms identified a putative neoblast-restricted gene set. This included many genes involved in chromatin modeling and RNA metabolism, suggesting that epigenetic modifications and post-transcriptional regulation are important for neoblast regulation

A mortalin-like gene is crucial for planarian stem cell viability

SUMMARY - In adult organisms, stem cells are crucial to homeostasis and regeneration of damaged tissues. In planarians, adult stem cells (neoblasts) are endowed with an extraordinary replicative potential that guarantees unlimited replacement of all differentiated cell types and extraordinary regenerative ability. The molecular mechanisms by which neoblasts combine long-term stability and constant proliferative activity, overcoming the impact of time, remain by far unknown. Here we investigate the role of Djmot, a planarian orthologue that encodes a peculiar member of the HSP70 family, named Mortalin, on the dynamics of stem cells of Dugesia japonica. Planarian stem cells and progenitors constitutively express Djmot. Transient Djmot expression in differentiated tissues is only observed after X-ray irradiation. DjmotRNA interference causes inability to regenerate and death of the animals, as a result of permanent growth arrest of stem cells. These results provide the first evidence that an hsp-related gene is essential for neoblast viability and suggest the possibility that high levels of Djmot serve to keep a p53-like protein signaling under control, thus allowing neoblasts to escape cell death programs. Further studies are needed to unravel the molecular pathways involved in these processes

Djeyes absent (Djeya) controls prototypic planarian eye regeneration by cooperating with the transcription factor Djsix-1

A conserved network of nuclear proteins is crucial to eye formation in both vertebrates and invertebrates. The finding that freshwater planarians can regenerate eyes without the contribution of Pax6 suggests that alternative combinations of regulatory elements may control the morphogenesis of the prototypic planarian eye. To further dissect the molecular events controlling eye regeneration in planarians, we investigated the role of eyes absent (Djeya) and six-1 (Djsix-1) genes in Dugesia japonica. These genes are expressed in both regenerating eyes and in differentiated photoreceptors of intact adults. Through RNAi studies, we show that Djsix-1 and Djeya are both critical for the regeneration of normal eyes in planarians and genetically cooperate in vivo to establish correct eye cell differentiation. We further demonstrate that the genetic interaction is mediated by physical interaction between the evolutionarily conserved domains of these two proteins. These data indicate that planarians use cooperatively Djsix-1 and Djeya for the proper specification of photoreceptors, implicating that the mechanism involving their evolutionarily conserved domains can be very ancient. Finally, both Djsix-1 and Djeya double-stranded RNA are substantially more effective at producing no-eye phenotypes in the second round of regeneration. This is probably due to the significant plasticity of the planarian model system, based on the presence of a stable population of totipotent stem cells, which ensure the rapid cell turnover of all differentiated cell types

SMC1A Codon 496 Mutations Affect the Cellular Response to Genotoxic Treatments

CorneliadeLangesyndromeisapleiotropicdevelopmentalsyndromecharacterizedbygrowthandcognitiveimpairment,facialdysmorphicfeatures,limbanomalies,andothermalfor-mations.MutationsincorecohesingenesSMC1AandSMC3,andthecohesinregulatorygene,NIPBL,havebeenidentifiedinCorneliadeLangesyndromeprobands.PatientswithNIPBLmutationshavemoreseverephenotypeswhencomparedtothosewithmutationsinSMC1AorSMC3.Todate,26distinctSMC1AmutationshavebeenidentifiedinpatientswithCorneliadeLangesyndrome.Here,wedescribea3-year-oldgirlwithpsy-chomotorandcognitiveimpairment,mildfacialdysmorphicfeaturesbutnolimbanomaly,heterozygousforac.1487G>AmutationinSMC1Awhichpredictsp.Arg496His.Weshowthatthismutationleadstoanimpairmentofthecellularresponsetogenotoxictreatments.2011WileyPeriodicals

Real-time PCR analysis of expression of 10 selected genes differentially regulated in 30 Gy irradiated planarians and controls

<p><b>Copyright information:</b></p><p>Taken from "Deciphering the molecular machinery of stem cells: a look at the neoblast gene expression profile"</p><p>Genome Biology 2007;8(4):R62-R62.</p><p>Published online 20 Apr 2007</p><p>PMCID:PMC1896013.</p><p></p> Expression levels are indicated in relative folds, assuming a value of 1 for untreated specimens (control). Values are expressed as mean ± standard deviation of six independent samples collected at each experimental condition conducted in duplicate. Genes are grouped in different charts according to the trend of their expression level in the analyzed samples. Plot analysis of the fold-change (30 Gy versus untreated controls) measured by real time reverse transcription (RT) polymerase chain reaction (RT-PCR) versus the fold change measured on the arrays. Values are represented in logarithmic scale. = 0.9851

Screenshot of Eisen's clustering based on 65 genes differentially regulated in 5 Gy and 30 Gy irradiated planarians and controls

<p><b>Copyright information:</b></p><p>Taken from "Deciphering the molecular machinery of stem cells: a look at the neoblast gene expression profile"</p><p>Genome Biology 2007;8(4):R62-R62.</p><p>Published online 20 Apr 2007</p><p>PMCID:PMC1896013.</p><p></p> The 65 genes are clustered by sample grouping (horizontal bars: cyan for controls, yellow for 5 Gy group, and orange for 30 Gy group). Clustering of experimental samples was performed according to the method of Eisen and coworkers [20]. Gene log2 ratios were average corrected across experimental samples and displayed according to the central method for display using a normalization factor, as recommended by Ross and coworkers [62]. The Tree-View software was used for the visualization. red, upregulated; green, downregulated; black, no difference

Graphical representation of the distribution (percentage) of genes by functional category

<p><b>Copyright information:</b></p><p>Taken from "Deciphering the molecular machinery of stem cells: a look at the neoblast gene expression profile"</p><p>Genome Biology 2007;8(4):R62-R62.</p><p>Published online 20 Apr 2007</p><p>PMCID:PMC1896013.</p><p></p> The Dj600 chip, genes that are downregulated as a consequence of high-dose X-ray treatment, and gene set that is upregulated as a consequence of low-dose X-ray treatment

Proteomic Profile Identifies Dysregulated Pathways in Cornelia de Lange Syndrome Cells with Distinct Mutations in <i>SMC1A</i> and <i>SMC3</i> Genes

Mutations in cohesin genes have been identified in Cornelia de Lange syndrome (CdLS), but its etiopathogenetic mechanisms are still poorly understood. To define biochemical pathways that are affected in CdLS, we analyzed the proteomic profile of CdLS cell lines carrying mutations in the core cohesin genes, <i>SMC1A</i> and <i>SMC3</i>. Dysregulated protein expression was found in CdLS probands compared to controls. The proteomics analysis was able to discriminate between probands harboring mutations in the different domains of the SMC proteins. In particular, proteins involved in the response to oxidative stress were specifically down-regulated in hinge mutated probands. In addition, the finding that CdLS cell lines show an increase in global oxidative stress argues that it could contribute to some CdLS phenotypic features such as premature physiological aging and genome instability. Finally, the <i>c-MYC</i> gene represents a convergent hub lying at the center of dysregulated pathways, and is down-regulated in CdLS. This study allowed us to highlight, for the first time, specific biochemical pathways that are affected in CdLS, providing plausible causal evidence for some of the phenotypic features seen in CdLS