Protein Kinase D regulates several aspects of development in Drosophila melanogaster

<p>Abstract</p> <p>Background</p> <p>Protein Kinase D (PKD) is an effector of diacylglycerol-regulated signaling pathways. Three isoforms are known in mammals that have been linked to diverse cellular functions including regulation of cell proliferation, differentiation, motility and secretory transport from the trans-Golgi network to the plasma membrane. In <it>Drosophila</it>, there is a single PKD orthologue, whose broad expression implicates a more general role in development.</p> <p>Results</p> <p>We have employed tissue specific overexpression of various PKD variants as well as tissue specific RNAi, in order to investigate the function of the PKD gene in <it>Drosophila</it>. Apart from a wild type (WT), a kinase dead (kd) and constitutively active (SE) <it>Drosophila </it>PKD variant, we also analyzed two human isoforms hPKD2 and hPKD3 for their capacity to substitute PKD activity in the fly. Overexpression of either WT or kd-PKD variants affected primarily wing vein development. However, overexpression of SE-PKD and PKD RNAi was deleterious. We observed tissue loss, wing defects and degeneration of the retina. The latter phenotype conforms to a role of PKD in the regulation of cytoskeletal dynamics. Strongest phenotypes were larval to pupal lethality. RNAi induced phenotypes could be rescued by a concurrent overexpression of <it>Drosophila </it>wild type PKD or either human isoform hPKD2 and hPKD3.</p> <p>Conclusion</p> <p>Our data confirm the hypothesis that <it>Drosophila </it>PKD is a multifunctional kinase involved in diverse processes such as regulation of the cytoskeleton, cell proliferation and death as well as differentiation of various fly tissues.</p

A Novel Pzg-NURF Complex Regulates Notch Target Gene Activity

The Putzig (Pzg) protein is associated with the NURF nucleosome remodeling complex, thereby promoting Notch target gene expression. Our findings suggest a novel Pzg-NURF complex that is responsible for the epigenetic regulation of Notch target genes

Linking aberrant chromatin features in chronic lymphocytic leukemia to transcription factor networks

In chronic lymphocytic leukemia (CLL), a diverse set of genetic mutations is embedded in a deregulated epigenetic landscape that drives cancerogenesis. To elucidate the role of aberrant chromatin features, we mapped DNA methylation, seven histone modifications, nucleosome positions, chromatin accessibility, binding of EBF1 and CTCF, as well as the transcriptome of B cells from CLL patients and healthy donors. A globally increased histone deacetylase activity was detected and half of the genome comprised transcriptionally downregulated partially DNA methylated domains demarcated by CTCF. CLL samples displayed a H3K4me3 redistribution and nucleosome gain at promoters as well as changes of enhancer activity and enhancer linkage to target genes. A DNA binding motif analysis identified transcription factors that gained or lost binding in CLL at sites with aberrant chromatin features. These findings were integrated into a gene regulatory enhancer containing network enriched for B‐cell receptor signaling pathway components. Our study predicts novel molecular links to targets of CLL therapies and provides a valuable resource for further studies on the epigenetic contribution to the disease

Large expert-curated database for benchmarking document similarity detection in biomedical literature search

Document recommendation systems for locating relevant literature have mostly relied on methods developed a decade ago. This is largely due to the lack of a large offline gold-standard benchmark of relevant documents that cover a variety of research fields such that newly developed literature search techniques can be compared, improved and translated into practice. To overcome this bottleneck, we have established the RElevant LIterature SearcH consortium consisting of more than 1500 scientists from 84 countries, who have collectively annotated the relevance of over 180 000 PubMed-listed articles with regard to their respective seed (input) article/s. The majority of annotations were contributed by highly experienced, original authors of the seed articles. The collected data cover 76% of all unique PubMed Medical Subject Headings descriptors. No systematic biases were observed across different experience levels, research fields or time spent on annotations. More importantly, annotations of the same document pairs contributed by different scientists were highly concordant. We further show that the three representative baseline methods used to generate recommended articles for evaluation (Okapi Best Matching 25, Term Frequency-Inverse Document Frequency and PubMed Related Articles) had similar overall performances. Additionally, we found that these methods each tend to produce distinct collections of recommended articles, suggesting that a hybrid method may be required to completely capture all relevant articles. The established database server located at https://relishdb.ict.griffith.edu.au is freely available for the downloading of annotation data and the blind testing of new methods. We expect that this benchmark will be useful for stimulating the development of new powerful techniques for title and title/abstract-based search engines for relevant articles in biomedical research.Peer reviewe

Loss of putzig Activity Results in Apoptosis during Wing Imaginal Development in Drosophila.

The Drosophila gene putzig (pzg) encodes a nuclear protein that is an integral component of the Trf2/Dref complex involved in the transcription of proliferation-related genes. Moreover, Pzg is found in a complex together with the nucleosome remodeling factor NURF, where it promotes Notch target gene activation. Here we show that downregulation of pzg activity in the developing wing imaginal discs induces an apoptotic response, accompanied by the induction of the pro-apoptotic gene reaper, repression of Drosophila inhibitor of apoptosis protein accumulation and the activation of the caspases Drice, Caspase3 and Dcp1. As a further consequence 'Apoptosis induced Proliferation' (AiP) and 'Apoptosis induced Apoptosis' (AiA) are triggered. As expected, the activity of the stress kinase Jun N-terminal kinase (JNK), proposed to mediate both processes, is ectopically induced in response to pzg loss. In addition, the expression of the mitogen wingless (wg) but not of decapentaplegic (dpp) is observed. We present evidence that downregulation of Notch activates Dcp1 caspase and JNK signaling, however, neither induces ectopic wg nor dpp expression. In contrast, the consequences of Dref-RNAi were largely indistinguishable from pzg-RNAi with regard to apoptosis induction. Moreover, overexpression of Dref ameliorated the downregulation of pzg compatible with the notion that the two are required together to maintain cell and tissue homeostasis in Drosophila

<i>pzg</i> depletion autonomously triggers the apoptotic signaling cascade in wing imaginal discs.

<p>RNAi mediated depletion of <i>pzg</i> was induced in the posterior part of the wing disc using <i>en</i>-Gal4. <i>Rpr</i>-lacZ, DIAP1 and caspase activity (in red) was monitored as indicated. <b>(A-A''')</b> A strong activation of the pro-apoptotic gene <i>rpr</i> (arrows) as well as the <i>Drosophila</i> activated caspases Drice^act (<b>C-C'''</b>, arrow), Caspase 3^act (<b>D-D'''</b>, arrow) and Dcp-1^act (<b>E-E'''</b>, arrows<i>)</i> is detected in the posterior half of the disc, whereas DIAP1 protein level is reduced (<b>B-B'''</b>, repressive arrow). (A-A''') <i>en-</i>Gal4 UAS-<i>GFP</i> UAS<i>-pzg-</i>RNAi/<i>+</i>; <i>rpr-</i>lacZ<i>/+</i>. (B-E''') <i>en-</i>Gal4 UAS-<i>GFP</i> UAS<i>-pzg-</i>RNAi/<i>+</i>. Pzg protein is shown in blue (anti-Pzg, A-E and A'''- E'''); GFP in green (<i>en-Gal4 GFP</i>) marks the posterior compartment (A-E and A''-E''). Posterior is to the right and dorsal up. The antero-posterior compartment boundary is marked with a dashed line. Scale bars: 100 μm.</p

<i>pzg</i>-RNAi induces AiA in late larval stages.

<p><b>(A-D'')</b> Strong caspase activity, either visualized with anti-Caspase 3^act (red in A-A', B-B') or anti-Dcp-1^act (red in C-C', D-D'), can be detected non-autonomously in late larval wing disc in the anterior compartment (arrowheads in A-D'). Preventing cell death execution with p35 amplifies the overgrowth effect (asterisks in B'') but still induces AiA in the anterior half of the disc (B-B'', D-D'', arrowheads). Genotypes: (A-A'') and (C-C'') <i>en-</i>Gal4 UAS-<i>GFP UAS-pzg-</i>RNAi/+. (B-B'') and (D-D'') UAS-<i>p35; en-</i>Gal4 UAS-<i>GFP UAS-pzg-</i>RNAi/+. The A/P compartment boundary is marked with a dotted line. Scale bars: 100 μm.</p

<i>pzg</i> genetically interacts with pro-apoptotic genes.

<p>(<b>A-C</b>) Control eyes after overexpression of <i>lacZ</i> and <i>pzg</i>, as well as <i>pzg-</i>RNAi induction. (<b>D-F</b>) Overexpression of <i>hid</i>, <i>rpr</i> and <i>grim</i> during eye development causes small and rough eyes in the adults. (<b>G-I</b>) Overexpression of <i>pzg</i> ameliorates the small eye size resulting from ectopically expressed pro-apoptotic genes, whereas depletion of <i>pzg</i> activity by RNAi enhances the phenotypes (<b>J-L</b>). Genotypes analyzed: <i>Gmr</i>-Gal4/+; UAS-<i>lacZ/+</i>. <i>Gmr</i>-Gal4/+; EP-<i>pzg</i>/+. <i>Gmr</i>-Gal4/+; UAS-<i>pzg-</i>RNAi/<i>+</i>. <i>Gmr-hid</i>/<i>Gmr</i>-Gal4; UAS-<i>lacZ</i>/+. <i>Gmr</i>-Gal4/+; <i>Gmr-rpr</i> or <i>Gmr-grim/</i> UAS-<i>lacZ</i>. <i>Gmr-hid</i>/<i>Gmr</i>-Gal4; EP-<i>pzg</i>/+. <i>Gmr</i>-Gal4/+; <i>Gmr-rpr</i> or <i>Gmr-grim</i>/EP-pzg. <i>Gmr-hid</i>/<i>Gmr</i>-Gal4; UAS-<i>pzg-</i>RNAi/<i>+</i>. <i>Gmr</i>-Gal4/+; <i>Gmr-rpr</i> or <i>Gmr-grim</i>/ UAS-<i>pzg-</i>RNAi.</p