Mutations in the Polycomb Group Gene polyhomeotic Lead to Epithelial Instability in both the Ovary and Wing Imaginal Disc in Drosophila

Most human cancers originate from epithelial tissues and cell polarity and adhesion defects can lead to metastasis. The Polycomb-Group of chromatin factors were first characterized in Drosophila as repressors of homeotic genes during development, while studies in mammals indicate a conserved role in body plan organization, as well as an implication in other processes such as stem cell maintenance, cell proliferation, and tumorigenesis. We have analyzed the function of the Drosophila Polycomb-Group gene polyhomeotic in epithelial cells of two different organs, the ovary and the wing imaginal disc.Clonal analysis of loss and gain of function of polyhomeotic resulted in segregation between mutant and wild-type cells in both the follicular and wing imaginal disc epithelia, without excessive cell proliferation. Both basal and apical expulsion of mutant cells was observed, the former characterized by specific reorganization of cell adhesion and polarity proteins, the latter by complete cytoplasmic diffusion of these proteins. Among several candidate target genes tested, only the homeotic gene Abdominal-B was a target of PH in both ovarian and wing disc cells. Although overexpression of Abdominal-B was sufficient to cause cell segregation in the wing disc, epistatic analysis indicated that the presence of Abdominal-B is not necessary for expulsion of polyhomeotic mutant epithelial cells suggesting that additional polyhomeotic targets are implicated in this phenomenon.Our results indicate that polyhomeotic mutations have a direct effect on epithelial integrity that can be uncoupled from overproliferation. We show that cells in an epithelium expressing different levels of polyhomeotic sort out indicating differential adhesive properties between the cell populations. Interestingly, we found distinct modalities between apical and basal expulsion of ph mutant cells and further studies of this phenomenon should allow parallels to be made with the modified adhesive and polarity properties of different types of epithelial tumors

The Elongin Complex Antagonizes the Chromatin Factor Corto for Vein versus Intervein Cell Identity in Drosophila Wings

International audienceDrosophila wings mainly consist of two cell types, vein and intervein cells. Acquisition of either fate depends on specific expression of genes that are controlled by several signaling pathways. The nuclear mechanisms that translate signaling into regulation of gene expression are not completely understood, but they involve chromatin factors from the Trithorax (TrxG) and Enhancers of Trithorax and Polycomb (ETP) families. One of these is the ETP Corto that participates in intervein fate through interaction with the Drosophila EGF Receptor - MAP kinase ERK pathway. Precise mechanisms and molecular targets of Corto in this process are not known. We show here that Corto interacts with the Elongin transcription elongation complex. This complex, that consists of three subunits (Elongin A, B, C), increases RNA polymerase II elongation rate in vitro by suppressing transient pausing. Analysis of phenotypes induced by EloA, B, or C deregulation as well as genetic interactions suggest that the Elongin complex might participate in vein vs intervein specification, and antagonizes corto as well as several TrxG genes in this process. Chromatin immunoprecipitation experiments indicate that Elongin C and Corto bind the vein-promoting gene rhomboid in wing imaginal discs. We propose that Corto and the Elongin complex participate together in vein vs intervein fate, possibly through tissue-specific transcriptional regulation of rhomboid

Drosophila Cyclin G and epigenetic maintenance of gene expression during development

Background: Cyclins and cyclin-dependent kinases (CDKs) are essential for cell cycle regulation and are functionally associated with proteins involved in epigenetic maintenance of transcriptional patterns in various developmental or cellular contexts. Epigenetic maintenance of transcription patterns, notably of Hox genes, requires the conserved Polycomb-group (PcG), Trithorax-group (TrxG), and Enhancer of Trithorax and Polycomb (ETP) proteins, particularly well studied in Drosophila. These proteins form large multimeric complexes that bind chromatin and appose or recognize histone post-translational modifications. PcG genes act as repressors, counteracted by trxG genes that maintain gene activation, while ETPs interact with both, behaving alternatively as repressors or activators. Drosophila Cyclin G negatively regulates cell growth and cell cycle progression, binds and co-localizes with the ETP Corto on chromatin, and participates with Corto in Abdominal-B Hox gene regulation. Here, we address further implications of Cyclin G in epigenetic maintenance of gene expression. Results: We show that Cyclin G physically interacts and extensively co-localizes on chromatin with the conserved ETP Additional sex combs (ASX), belonging to the repressive PR-DUB complex that participates in H2A deubiquitination and Hox gene silencing. Furthermore, Cyclin G mainly co-localizes with RNA polymerase II phosphorylated on serine 2 that is specific to productive transcription. CycG interacts with Asx, PcG, and trxG genes in Hox gene maintenance, and behaves as a PcG gene. These interactions correlate with modified ectopic Hox protein domains in imaginal discs, consistent with a role for Cyclin G in PcG-mediated Hox gene repression. Conclusions: We show here that Drosophila CycG is a Polycomb-group gene enhancer, acting in epigenetic maintenance of the Hox genes Sex combs reduced (Scr) and Ultrabithorax (Ubx). However, our data suggest that Cyclin G acts alternatively as a transcriptional activator or repressor depending on the developmental stage, the tissue or the target gene. Interestingly, since Cyclin G interacts with several CDKs, Cyclin G binding to the ETPs ASX or Corto suggests that their activity could depend on Cyclin G-mediated phosphorylation. We discuss whether Cyclin G fine-tunes transcription by controlling H2A ubiquitination and transcriptional elongation via interaction with the ASX subunit of PR-DUB.Science, Faculty ofZoology, Department ofReviewedFacult

Can pre-implantation biopsies predict renal allograft function in paediatric renal transplant recipients ?

<p>The upper allele was brought by the mother. The number of females with ectopic veins among flies transheterozygous for <i>Elo</i> and <i>corto</i> mutations was compared to the number of females with ectopic veins among flies with a <i>corto</i> mutation only (z-test, ^a p<0.001).</p

<i>Elo</i> genes control wing cell identity.

<p>(A): Wing from control <i>w¹¹¹⁸</i> fly (L1-L5: longitudinal veins; ACV and PCV: anterior and posterior cross-veins). (B, C): Wings from <i>+/EloB^EP3132</i> and <i>EloB^EP3132/Df(3R)BSC518</i> flies exhibit truncated L5. (D): Wings from <i>+/sd::Gal4</i> flies have a very faint ectopic vein phenotype and no margin phenotype. (E, F): Wings from flies over-expressing <i>EloA</i> exhibit ectopic vein and margin phenotypes. (G, H, I): <i>EloC^SH1520</i> and <i>EloC^SH1299</i> loss-of-function alleles diminish expressivity of the ectopic vein phenotype induced by the <i>bs^EY23316</i> loss-of-function allele. Strong phenotype: ectopic veins everywhere in the wing (shown in G). Mild phenotype: ectopic veins under the posterior cross-vein only (shown in H).</p

Decreasing <i>EloC</i> expression suppresses ectopic veins induced by <i>blistered</i> loss-of-function.

<p>The upper allele was brought by the mother. The number of <i>EloC</i>/<i>bs^EY23316</i> females with ectopic veins was compared to the number of <i>+/bs^EY23316</i> females with ectopic veins (z-test, ^a p<0.001). The mild ectopic vein phenotype corresponds to presence of ectopic veins distal to the posterior cross-vein (<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0077592#pone-0077592-g005" target="_blank">Figure 5H</a>), whereas the strong ectopic vein phenotype corresponds to presence of ectopic veins everywhere in the wing (<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0077592#pone-0077592-g005" target="_blank">Figure 5G</a>).</p

Down-regulation of <i>EloC</i> by RNA interference impairs both cell proliferation and cell differentiation in wing imaginal discs.

<p>(A): Clones expressing the <i>ValEloC</i> transgene (GFP⁺ cells, shown by white arrows) are located at the periphery of the disc and are very small compared to control clones. (B, C, D): Wings from pharates in which <i>ValEloC</i> is driven by <i>nub::Gal4</i> (C) or <i>rn::Gal4</i> (D), both expressed in the wing pouch <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0077592#pone.0077592-StPierre1" target="_blank">[66]</a>, <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0077592#pone.0077592-Ng1" target="_blank">[67]</a> are small compared to wild-type pharate wings (B) and exhibit severe wing blade defects. By contrast, longitudinal veins (shown by asterisks) are formed in the proximal-most part of the wing blade where <i>nub::Gal4</i> and <i>rn::Gal4</i> are not expressed.</p

<i>corto</i> and several TrxG genes control wing cell identity.

<p>(A, B): Ectopic vein phenotypes induced by the <i>corto⁰⁷¹²⁸</i> loss-of-function allele (A) or by <i>corto⁴²⁰</i> loss-of-function clones (B). (C): <i>corto⁴²⁰</i> homozygous clones (GFP^- cells) in wing imaginal discs. (D, E, F): Ectopic vein phenotypes induced by <i>mor</i>, <i>kis</i> or <i>trx</i> loss-of-function alleles. In A, B, D, E, F, asterisks mark ectopic veins.</p