The histone‐binding protein COPR5 is required for nuclear functions of the protein arginine methyltransferase PRMT5Lacroix, Matthieu; Messaoudi, Selma El; Rodier, Geneviève; Le Cam, Aphonse; Sardet, Claude; Fabbrizio, Eric
doi: 10.1038/embor.2008.45pmid: 18404153
Protein arginine methyltransferase 5 (PRMT5) targets nuclear and cytoplasmic proteins. Here, we identified a nuclear protein, called cooperator of PRMT5 (COPR5), involved in the nuclear functions of PRMT5. COPR5 tightly binds to PRMT5, both in vitro and in living cells, but not to other members of the PRMT family. PRMT5 bound to COPR5 methylates histone H4 (R3) preferentially when compared with histone H3 (R8), suggesting that COPR5 modulates the substrate specificity of nuclear PRMT5‐containing complexes, at least towards histones. Markedly, recombinant COPR5 binds to the amino terminus of histone H4 and is required to recruit PRMT5 to reconstituted nucleosomes in vitro. Consistently, COPR5 depletion in cells strongly reduces PRMT5 recruitment on chromatin at the PRMT5 target gene cyclin E1 (CCNE1) in vivo. Moreover, both COPR5 depletion and overexpression affect CCNE1 promoter expression. We propose that COPR5 is an important chromatin adaptor for PRMT5 to function on a subset of its target genes.
Gene induction following wounding of wild‐type versus macrophage‐deficient Drosophila embryosStramer, Brian; Winfield, Mark; Shaw, Tanya; Millard, Thomas H; Woolner, Sarah; Martin, Paul
doi: 10.1038/embor.2008.34pmid: 18344972
By using a microarray screen to compare gene responses after sterile laser wounding of wild‐type and ‘macrophageless’ serpent mutant Drosophila embryos, we show the wound‐induced programmes that are independent of a pathogenic response and distinguish which of the genes are macrophage dependent. The evolutionarily conserved nature of this response is highlighted by our finding that one such new inflammation‐associated gene, growth arrest and DNA damage‐inducible gene 45 (GADD45), is upregulated in both Drosophila and murine repair models. Comparison of unwounded wild‐type and serpent mutant embryos also shows a portfolio of ‘macrophage‐specific’ genes, which suggest analogous functions with vertebrate inflammatory cells. Besides identifying the various classes of wound‐ and macrophage‐related genes, our data indicate that sterile injury per se, in the absence of pathogens, triggers induction of a ‘pathogen response’, which might prime the organism for what is likely to be an increased risk of infection.
No single way to understand singlet oxygen signalling in plantsKim, Chanhong; Meskauskiene, Rasa; Apel, Klaus; Laloi, Christophe
doi: 10.1038/embor.2008.57pmid: 18451767
When plant cells are under environmental stress, several chemically distinct reactive oxygen species (ROS) are generated simultaneously in various intracellular compartments and these can cause oxidative damage or act as signals. The conditional flu mutant of Arabidopsis, which generates singlet oxygen in plastids during a dark‐to‐light transition, has allowed the biological activity of singlet oxygen to be determined, and the criteria to distinguish between cytotoxicity and signalling of this particular ROS to be defined. The genetic basis of singlet‐oxygen‐mediated signalling has been revealed by the mutation of two nuclear genes encoding the plastid proteins EXECUTER (EX)1 and EX2, which are sufficient to abrogate singlet‐oxygen‐dependent stress responses. Conversely, responses due to higher cytotoxic levels of singlet oxygen are not suppressed in the ex1/ex2 background. Whether singlet oxygen levels lower than those that trigger genetically controlled cell death activate acclimation is now under investigation.
Substrate‐induced DNA strand misalignment during catalytic cycling by DNA polymerase λBebenek, Katarzyna; Garcia‐Diaz, Miguel; Foley, Meredith C; Pedersen, Lars C; Schlick, Tamar; Kunkel, Thomas A
doi: 10.1038/embor.2008.33pmid: 18369368
The simple deletion of nucleotides is common in many organisms. It can be advantageous when it activates genes beneficial to microbial survival in adverse environments, and deleterious when it mutates genes relevant to survival, cancer or degenerative diseases. The classical idea is that simple deletions arise by strand slippage. A prime opportunity for slippage occurs during DNA synthesis, but it remains unclear how slippage is controlled during a polymerization cycle. Here, we report crystal structures and molecular dynamics simulations of mutant derivatives of DNA polymerase λ bound to a primer–template during strand slippage. Relative to the primer strand, the template strand is in multiple conformations, indicating intermediates on the pathway to deletion mutagenesis. Consistent with these intermediates, the mutant polymerases generate single‐base deletions at high rates. The results indicate that dNTP‐induced template strand repositioning during conformational rearrangements in the catalytic cycle is crucial to controlling the rate of strand slippage.
Formation of a new receptor‐operated channel by heteromeric assembly of TRPP2 and TRPC1 subunitsBai, Chang‐Xi; Giamarchi, Aurélie; Rodat‐Despoix, Lise; Padilla, Françoise; Downs, Tamyra; Tsiokas, Leonidas; Delmas, Patrick
doi: 10.1038/embor.2008.29pmid: 18323855
Although several protein–protein interactions have been reported between transient receptor potential (TRP) channels, they are all known to occur exclusively between members of the same group. The only intergroup interaction described so far is that of TRPP2 and TRPC1; however, the significance of this interaction is unknown. Here, we show that TRPP2 and TRPC1 assemble to form a channel with a unique constellation of new and TRPP2/TRPC1‐specific properties. TRPP2/TRPC1 is activated in response to G‐protein‐coupled receptor activation and shows a pattern of single‐channel conductance, amiloride sensitivity and ion permeability distinct from that of TRPP2 or TRPC1 alone. Native TRPP2/TRPC1 activity is shown in kidney cells by complementary gain‐of‐function and loss‐of‐function experiments, and its existence under physiological conditions is supported by colocalization at the primary cilium and by co‐immunoprecipitation from kidney membranes. Identification of the heteromultimeric TRPP2/TRPC1 channel has implications in mechanosensation and cilium‐based Ca2+ signalling.
‘Insulator bodies’ are aggregates of proteins but not of insulatorsGolovnin, Anton; Melnikova, Larisa; Volkov, Ilya; Kostuchenko, Margarita; Galkin, Alexander V; Georgiev, Pavel
doi: 10.1038/embor.2008.32pmid: 18369369
Chromatin insulators are thought to restrict the action of enhancers and silencers. The best‐known insulators in Drosophila require proteins such as Suppressor of Hairy wing (Su(Hw)) and Modifier of mdg4 (Mod(mdg4)) to be functional. The insulator‐related proteins apparently colocalize as nuclear speckles in immunostained cells. It has been asserted that these speckles are ‘insulator bodies’ of many Su(Hw)–insulator DNA sites held together by associated proteins, including Mod(mdg4). As we show here using flies, larvae and S2 cells, a mutant Mod(mdg4) protein devoid of the Q‐rich domain supports the function of Su(Hw)‐dependent insulators and efficiently binds to correct insulator sites on the chromosome, but does not form or enter the Su(Hw)‐marked nuclear speckles; conversely, the latter accumulate another (C‐truncated) Mod(mdg4) mutant that cannot interact with Su(Hw) or with the genuine insulators. Hence, it is not the functional genomic insulators but rather aggregated proteins that make the so‐called ‘insulator bodies’.