Non-structural protein 1 (NS1) of influenza virus has been shown to inhibit the innate immune response by blocking the induction of interferon (IFN). In this study, we isolated two single-stranded RNA aptamers specific to NS1 with K d values of 1.62 ± 0.30 nM and 1.97 ± 0.27 nM, respectively, using a systematic evolution of ligand by exponential enrichment (SELEX) procedure. The selected aptamers were able to inhibit the interaction of NS1 with tripartite motif-containing protein 25 (TRIM25), and suppression of NS1 enabled retinoic acid inducible gene I (RIG-I) to be ubiquitinated regularly by TRIM25. Additional luciferase reporter assay and quantitative real-time PCR (RT-PCR) experiments demonstrated that suppression of NS1 by the selected aptamers induced IFN production. It is noted that viral replication was also inhibited through IFN induction in the presence of the selected aptamers. These results suggest that the isolated aptamers are strongly expected to be new therapeutic agents against influenza infection.
Monoubiquitination of histone H2B, catalyzed by Rad6-Bre1, is required for methylation of histone H3 on lysines 4 and 79, catalyzed by the Set1-containing complex COMPASS and Dot1p, respectively. The Paf1 protein complex, which associates with RNA polymerase II, is known to be required for these histone H3 methylation events. During the early elongation stage of transcription, the Paf1 complex is required for association of COMPASS with RNA polymerase II, but the role the Paf1 complex plays at the promoter has not been clear. We present evidence that the Paf1 complex is required for monoubiquitination of histone H2B at promoters. Strains deleted for several components of the Paf1 complex are defective in monoubiquitination of histone H2B, which results in the loss of methylation of lysines 4 and 79 of histone H3. We also show that Paf1 complex is required for the interaction of Rad6 and COMPASS with RNA polymerase II. Finally, we show that the Paf1 complex is required for Rad6-Bre1 catalytic activity but not for the recruitment of Rad6-Bre1 to promoters. Thus, in addition to its role during the elongation phase of transcription, the Paf1 complex appears to activate the function but not the placement of the Rad6-Bre1 ubiquitin-protein ligase at the promoters of active genes. Monoubiquitination of histone H2B, catalyzed by Rad6-Bre1, is required for methylation of histone H3 on lysines 4 and 79, catalyzed by the Set1-containing complex COMPASS and Dot1p, respectively. The Paf1 protein complex, which associates with RNA polymerase II, is known to be required for these histone H3 methylation events. During the early elongation stage of transcription, the Paf1 complex is required for association of COMPASS with RNA polymerase II, but the role the Paf1 complex plays at the promoter has not been clear. We present evidence that the Paf1 complex is required for monoubiquitination of histone H2B at promoters. Strains deleted for several components of the Paf1 complex are defective in monoubiquitination of histone H2B, which results in the loss of methylation of lysines 4 and 79 of histone H3. We also show that Paf1 complex is required for the interaction of Rad6 and COMPASS with RNA polymerase II. Finally, we show that the Paf1 complex is required for Rad6-Bre1 catalytic activity but not for the recruitment of Rad6-Bre1 to promoters. Thus, in addition to its role during the elongation phase of transcription, the Paf1 complex appears to activate the function but not the placement of the Rad6-Bre1 ubiquitin-protein ligase at the promoters of active genes. The DNA of eukaryotic organisms assembles around histone proteins in nucleosomes to form highly organized structures known as chromatin (1Berger S.L. Curr. Opin. Genet. Dev. 2002; 12: 142-148Crossref PubMed Scopus (991) Google Scholar). Alterations in chromatin structure play a major role in regulating gene expression, and for this reason much attention has been focused recently on the covalent modifications of histone proteins and their outcomes in transcriptional elongation (1Berger S.L. Curr. Opin. Genet. Dev. 2002; 12: 142-148Crossref PubMed Scopus (991) Google Scholar, 2Gerber M. Shilatifard A. J. Biol. Chem. 2003; 278: 26303-26306Abstract Full Text Full Text PDF PubMed Scopus (141) Google Scholar, 3Kouzarides T. Curr. Opin. Genet. Dev. 2002; 12: 198-209Crossref PubMed Scopus (748) Google Scholar, 4Turner B.M. Cell. 2002; 111: 285-291Abstract Full Text Full Text PDF PubMed Scopus (930) Google Scholar). Essential to this process are the N-terminal tails of histone proteins. Because they protrude from the globular body of the nucleosome and are available for interactions with other proteins, the tails are the site of many covalent modifications that alter nucleosome structure. A myriad of modifications, such as acetylation, phosphorylation, ubiquitination, and methylation, decorate each histone tail (2Gerber M. Shilatifard A. J. Biol. Chem. 2003; 278: 26303-26306Abstract Full Text Full Text PDF PubMed Scopus (141) Google Scholar, 5Jenuwein T. Allis C.D. Science. 2001; 293: 1074-1080Crossref PubMed Scopus (7709) Google Scholar) The combinatorial effects of such modifications can produce an array of different responses involved in transcriptional activation and repression (1Berger S.L. Curr. Opin. Genet. Dev. 2002; 12: 142-148Crossref PubMed Scopus (991) Google Scholar, 2Gerber M. Shilatifard A. J. Biol. Chem. 2003; 278: 26303-26306Abstract Full Text Full Text PDF PubMed Scopus (141) Google Scholar, 3Kouzarides T. Curr. Opin. Genet. Dev. 2002; 12: 198-209Crossref PubMed Scopus (748) Google Scholar, 4Turner B.M. Cell. 2002; 111: 285-291Abstract Full Text Full Text PDF PubMed Scopus (930) Google Scholar, 5Jenuwein T. Allis C.D. Science. 2001; 293: 1074-1080Crossref PubMed Scopus (7709) Google Scholar, 6Jaskelioff M. Peterson C.L. Nat. Cell Biol. 2003; 5: 395-399Crossref PubMed Scopus (69) Google Scholar). One histone modification of major consequence is the methylation of histone H3 at lysines 4 and 79, catalyzed by the Set1-containing complex COMPASS 1The abbreviations used are: COMPASS, complex of proteins associated with Set1; Pol II, polymerase II; E3, ubiquitin-protein isopeptide ligase; ChIP, chromatin immunoprecipitation; TAP, tandem affinity purification; NIB, nuclear isolation buffer. and Dot1p, respectively (7Miller T. Krogan N.J. Dover J. Erdjument-Bromage H. Tempst P. Johnston M. Greenblatt J.F. Shilatifard A. Proc. Natl. Acad. Sci. U. S. A. 2001; 98: 12902-12907Crossref PubMed Scopus (464) Google Scholar, 8Krogan N.J. Dover J. Khorrami S. Greenblatt J.F. Schneider J. Johnston M. Shilatifard A. J. Biol. Chem. 2002; 277: 10753-10755Abstract Full Text Full Text PDF PubMed Scopus (322) Google Scholar, 9Nagy P.L. Griesenbeck J. Kornberg R.D. Cleary M.L. Proc. Natl. Acad. Sci. U. S. A. 2002; 99: 90-94Crossref PubMed Scopus (268) Google Scholar, 10Roguev A. Schaft D. Shevchenko A. Pijnappel W.W.M.P. Wilm M. Aasland R. Stewart A.F. EMBO J. 2001; 20: 7137-7148Crossref PubMed Scopus (461) Google Scholar, 11Ng H.H. Feng Q. Wang H. Erdjument-Bromage H. Tempst P. Zhang Y. Struhl K. Genes Dev. 2002; 16: 1518-1527Crossref PubMed Scopus (427) Google Scholar, 12Feng Q. Wang H. Ng H.H. Erdjument-Bromage H. Tempst P. Struhl K. Zhang Y. Cur. Biol. 2002; 12: 1052-1058Abstract Full Text Full Text PDF PubMed Scopus (646) Google Scholar, 13Briggs S.D. Bryk M. Strahl B.D. Cheung W.L. Davie J.K. Dent S.Y.R. Winston F. Allis C.D. Genes Dev. 2001; 15: 3286-3295Crossref PubMed Scopus (481) Google Scholar, 14van Leeuwen F. Gafken P.R. Gottschling D.E. Cell. 2002; 109: 745-756Abstract Full Text Full Text PDF PubMed Scopus (674) Google Scholar). It has been shown that methylation of both lysine residues impacts the expression of genes within the rDNA loci and telomeric regions of DNA in Saccharomyces cerevisiae (7Miller T. Krogan N.J. Dover J. Erdjument-Bromage H. Tempst P. Johnston M. Greenblatt J.F. Shilatifard A. Proc. Natl. Acad. Sci. U. S. A. 2001; 98: 12902-12907Crossref PubMed Scopus (464) Google Scholar, 8Krogan N.J. Dover J. Khorrami S. Greenblatt J.F. Schneider J. Johnston M. Shilatifard A. J. Biol. Chem. 2002; 277: 10753-10755Abstract Full Text Full Text PDF PubMed Scopus (322) Google Scholar, 15Bryk M. Briggs S.D. Strahl B.D. Curcio M.J. Allis C.D. Winston F. Curr. Biol. 2002; 12: 165-170Abstract Full Text Full Text PDF PubMed Scopus (173) Google Scholar, 16Ng H.H. Ciccone D.N. Morshead K.B. Oettinger M.A. Struhl K. Proc. Natl. Acad. Sci. U. S. A. 2003; 100: 1820-1825Crossref PubMed Scopus (250) Google Scholar). It has been demonstrated that some of the components of the Paf1 complex, a complex that associates with the initiating and elongating RNA polymerase II, is also required for histone H3 methylation on lysines 4 and 79 (17Krogan N.J. Dover J. Wood A. Schneider J. Heidt J. Boateng M.A. Dean K. Ryan O.W. Golshani A. Johnston M. Greenblatt J.F. Shilatifard A. Mol. Cell. 2003; 11: 721-729Abstract Full Text Full Text PDF PubMed Scopus (570) Google Scholar, 18Ng H.H. Robert F. Young R.A. Struhl K. Mol. Cell. 2003; 11: 709-719Abstract Full Text Full Text PDF PubMed Scopus (857) Google Scholar). Accordingly, previous studies have demonstrated a role for the Paf1 complex in transcriptional elongation and initiation (19Mueller C.L. Jaehning J.A. Mol. Cell. Biol. 2002; 22: 1971-1980Crossref PubMed Scopus (179) Google Scholar, 20Pokholok D.K. Hannett N.M. Young R.A. Mol. Cell. 2002; 9: 799-809Abstract Full Text Full Text PDF PubMed Scopus (266) Google Scholar, 21Porter S.E. Washburn T.M. Chang M. Jaehning J.A. Eukaryotic Cell. 2002; 1: 830-842Crossref PubMed Scopus (58) Google Scholar, 22Squazzo S.L. Costa P.J. Lindstrom D.L. Kumer K.E. Simic R. Jennings J.L. Link A.J. Arndt K.M. Hartzog G.A. EMBO J. 2002; 21: 1764-1774Crossref PubMed Scopus (247) Google Scholar). A further requirement for the methylation of both lysines 4 and 79 of histone H3 is the prior modification of another histone protein; monoubiquitination of histone H2B, catalyzed by the Rad6-Bre1 protein-ubiquitin ligase (23Robzyk K. Recht J. Osley M.A. Science. 2000; 287: 501-504Crossref PubMed Scopus (529) Google Scholar, 24Dover J. Schneider J. Tawiah-Boateng M.A. Wood A. Dean K. Johnston M. Shilatifard A. J. Biol. Chem. 2002; 277: 28368-28371Abstract Full Text Full Text PDF PubMed Scopus (428) Google Scholar, 25Sun Z.W. Allis C.D. Nature. 2002; 418: 104-108Crossref PubMed Scopus (838) Google Scholar, 26Wood A. Krogan N.J. Dover J. Schneider J. Heidt J. Boateng M.A. Dean K. Golshani A. Zhang Y. Greenblatt J.F. Johnston M. Shilatifard A. Mol. Cell. 2003; 11: 267-274Abstract Full Text Full Text PDF PubMed Scopus (410) Google Scholar, 27Ng H.H. Xu R.-M. Zhang Y. Struhl K. J. Biol. Chem. 2002; 277: 34655-34657Abstract Full Text Full Text PDF PubMed Scopus (340) Google Scholar, 28Briggs S.D. Xiao T. Sun Z.W. Caldwell J.A. Shabanowitz J. Hunt D.F. Allis C.D. Strahl B.D. Nature. 2002; 418: 498Crossref PubMed Scopus (395) Google Scholar). Although the Paf1 complex also plays a role in histone H3 Lys-36 methylation by Set2, this modification does not require monoubiquitination of histone H2B. Because the Paf1 complex is associated with RNA polymerase II at the promoter, we wished to determine whether it is also required for the monoubiquitination of histone H2B. Here we present evidence that components of the Paf1 complex are required for monoubiquitination of histone H2B. Strains in which several components of the Paf1 complex are deleted are defective in monoubiquitination of histone H2B at promoter regions, and this in turn results in the loss of methylation on histone H3 on lysines 4 and 79 within the body of active genes. Previously we demonstrated that the Paf1 complex is required for the interaction between RNA polymerase II and COMPASS, and that loss of the Paf1 subunit Ctr9 dissolved this interaction (17Krogan N.J. Dover J. Wood A. Schneider J. Heidt J. Boateng M.A. Dean K. Ryan O.W. Golshani A. Johnston M. Greenblatt J.F. Shilatifard A. Mol. Cell. 2003; 11: 721-729Abstract Full Text Full Text PDF PubMed Scopus (570) Google Scholar). We have recently found that Rad6 associates with RNA Pol II and COMPASS, and the Paf1 complex mediates this interaction as well. However, chromatin immunoprecipitation (ChIP) experiments indicate that the Paf1 complex is not required for the recruitment of Rad6-Bre1 to the promoter. Together, our studies indicate a role for the Paf1 complex at the promoter in regulating the functional activity of the Rad6-Bre1 complex in monoubiquitination of histone H2B, which is distinct from the role of the Paf1 complex in histone methylation during the process of transcription elongation. Preparation of Histone Extracts—Histone extraction was performed as described in (31Edmondson D.G. Smith M.M. Roth S.Y. Genes Dev. 1996; 10: 1247-1259Crossref PubMed Scopus (407) Google Scholar) with several changes. 50 ml of YPD (yeast/peptone/dextrose) were inoculated and allowed to grow at 30 °Ctoan A 600 of ∼1. The cultures were then transferred to 50-ml conical tubes and the cells pelleted by centrifugation at 2000 rpm. Cells were then resuspended in 500 μl of sterile water and transferred to 1.5-ml Eppendorf tubes (cell pellet size ∼150–200 μl). Cells were then pelleted by centrifugation at 2000 rpm, resuspended in 500 μl of NIB (0.25 m sucrose, 60 mm KCl, 14 mm NaCl, 5 mm MgCl2, 1 mm CaCl2, 0.8% Triton X-100), and then pelleted again. After removal of the supernatant by pipetting, cells were resuspended in another 500 μl of NIB, and ∼150 μl of 0.5-mm glass beads were added to one-third of the total volume of the suspension. The tubes were then closed, and lids were wrapped in parafilm to prevent leakage during lysis. The tubes were then vortexed vigorously for 15 min at 4 °C, and lysates were recovered into 15-ml conical tubes by puncturing the bottom of the 1.5-ml tube with a small-gauge needle, inserting the punctured tube into the top of the 15-ml tube, and briefly centrifuging the contents at 2000 rpm. The supernatants were discarded, and cell pellets were washed with 1 ml of chilled NIB followed by centrifugation to pellet the nuclei. After the supernatant was discarded, the nuclei were suspended in 200 μl of 0.4 n H2SO4 and incubated on ice for 30 min with occasional vortexing. Cellular debris were then pelleted by centrifugation at 2000 rpm. The supernatants were transferred to a fresh 1.5-ml tube, acetone was added at a 1:5 ratio (1 ml in this case), and the tubes were kept on ice for 45 min. Free histones were pelleted by centrifugation at 15,000 rpm, the supernatants were drained, and the pellets were resuspended in 75 μl of sterile water and 25 μl of Laemmli loading buffer. Extracts were then heated at 95 °C for 5 min and resolved using 16% SDS-PAGE. ChIP Experiments—Overnight culture of cells were treated with formaldehyde to achieve cross-linking as described previously (26Wood A. Krogan N.J. Dover J. Schneider J. Heidt J. Boateng M.A. Dean K. Golshani A. Zhang Y. Greenblatt J.F. Johnston M. Shilatifard A. Mol. Cell. 2003; 11: 267-274Abstract Full Text Full Text PDF PubMed Scopus (410) Google Scholar). After the cross-linked cells were lysed, the chromatin was isolated and sheared by sonication. The sheared chromatin solution was then subjected to immunoprecipitation using IgG-Sepharose beads. Immunoprecipitates were eluted and de-cross-linked, and then the free DNA was precipitated and used for PCR with primer pairs directed against different regions of the PMA1 gene. Each reaction contains a control primer set directed against a region of yeast chromosome 5 with no open reading frames. The -fold enrichment was determined by comparing the ratio of the immunoprecipitated PCR product to input DNA PCR product using ImageQuant software (Amersham Biosciences). Bre1 Is Required for Recruitment of Rad6 to the Promoter— Rad6 is required for monoubiquitination of histone H2B on lysine 123 (23Robzyk K. Recht J. Osley M.A. Science. 2000; 287: 501-504Crossref PubMed Scopus (529) Google Scholar) and is recruited to promoters via Bre1, one of its E3 ubiquitin ligases (26Wood A. Krogan N.J. Dover J. Schneider J. Heidt J. Boateng M.A. Dean K. Golshani A. Zhang Y. Greenblatt J.F. Johnston M. Shilatifard A. Mol. Cell. 2003; 11: 267-274Abstract Full Text Full Text PDF PubMed Scopus (410) Google Scholar). We have demonstrated that Bre1 interacts with Rad6 through its C-terminal C3HC4 ring finger domain, and thus perturbation of this domain by the addition of the TAP-tag to the C terminus of Bre1 abolishes the interaction of Rad6 with Bre1. However, this TAP-tagged Bre1 is still capable of being targeted to the promoter, suggesting that (a) Bre1 is the protein that targets Rad6 to promoters and (b) the C-terminal domain of Bre1 is dispensable for its localization to promoters (Fig. 1). Paf1 Complex Affects Both Histone H3 Methylation and H2B Ubiquitination—Both the components of the Paf1 complex and the ubiquitination activity of the Rad6-Bre1 complex are required for methylation of histone H3 on both lysines 4 and 79 (Fig. 2A) (24Dover J. Schneider J. Tawiah-Boateng M.A. Wood A. Dean K. Johnston M. Shilatifard A. J. Biol. Chem. 2002; 277: 28368-28371Abstract Full Text Full Text PDF PubMed Scopus (428) Google Scholar, 25Sun Z.W. Allis C.D. Nature. 2002; 418: 104-108Crossref PubMed Scopus (838) Google Scholar, 26Wood A. Krogan N.J. Dover J. Schneider J. Heidt J. Boateng M.A. Dean K. Golshani A. Zhang Y. Greenblatt J.F. Johnston M. Shilatifard A. Mol. Cell. 2003; 11: 267-274Abstract Full Text Full Text PDF PubMed Scopus (410) Google Scholar, 27Ng H.H. Xu R.-M. Zhang Y. Struhl K. J. Biol. Chem. 2002; 277: 34655-34657Abstract Full Text Full Text PDF PubMed Scopus (340) Google Scholar). To determine whether the Paf1 complex is also required for histone H2B ubiquitination via Rad6-Bre1, we tested whether H2B is ubiquitinated in the absence of either of the Paf1 complex subunits Rtf1 or Paf1. Interestingly, deletion of either RTF1 or PAF1 resulted in the loss of histone H2B ubiquitination (Fig. 2B). Ubiquitination of histone H2B and methylation of histone H3 were complemented by introducing a plasmid containing either wild-type RTF1 or PAF1 into the mutant cells lacking Rtf1 or Paf1 (Fig. 2C). Interaction among Rad6, COMPASS, and RNA Polymerase II Requires Paf1 Complex—COMPASS interacts with RNA polymerase II during the early stages of transcriptional elongation (17Krogan N.J. Dover J. Wood A. Schneider J. Heidt J. Boateng M.A. Dean K. Ryan O.W. Golshani A. Johnston M. Greenblatt J.F. Shilatifard A. Mol. Cell. 2003; 11: 721-729Abstract Full Text Full Text PDF PubMed Scopus (570) Google Scholar, 18Ng H.H. Robert F. Young R.A. Struhl K. Mol. Cell. 2003; 11: 709-719Abstract Full Text Full Text PDF PubMed Scopus (857) Google Scholar). To determine whether there are interactions between COMPASS, RNA polymerase II, and Rad6, COMPASS was purified via one of its tagged subunits, the Ash2 homologue CPS60/Bre2. RNA polymerase II co-purifies with COMPASS, as does Rad6 (Fig. 3, A–C, lanes 6 and 7), indicating that a functional as well as physical interaction exists between the ubiquitination and methylation machinery at the promoter region of active genes. However, in strains missing the gene encoding the Rtf1 subunit of the Paf1 complex, Rad6 no longer associates with COMPASS and RNA polymerase II (Fig. 3C, lanes 8 and 9), but the interaction between COMPASS and Pol II remains stable (Fig. 3, A and B, lanes 8 and 9). From this it seems likely that the deletion of Rtf1 does not affect the stability of the Paf1 complex but may instead be a part of the site where Rad6 interacts. Previously, we demonstrated that the Ctr9 subunit (one of the largest subunit) of the Paf1 complex is required for the interaction of COMPASS and RNA polymerase II (17Krogan N.J. Dover J. Wood A. Schneider J. Heidt J. Boateng M.A. Dean K. Ryan O.W. Golshani A. Johnston M. Greenblatt J.F. Shilatifard A. Mol. Cell. 2003; 11: 721-729Abstract Full Text Full Text PDF PubMed Scopus (570) Google Scholar). However, removal of the Rtf1 component of the Paf1 complex has no effect on the interaction between RNA polymerase II and COMPASS (Fig. 3, A and B, lanes 8 and 9), indicating that COMPASS may physically interact with Ctr9 subunit of the Paf1 complex or that Ctr9 is essential for the stability of the Paf1 complex and therefore its interaction with RNA polymerase II. We have also demonstrated that the interaction between RNA polymerase II and COMPASS with Rad6 is dependent on the presence of the Rtf1 subunit of the Paf1 complex (Fig. 3C, lanes 6–9). Paf1 Complex Is Not Necessary for Rad6 Recruitment to the Promoter—A simple model that could account for these observations is that the Paf1 complex is responsible for the localization of the Rad6-Bre1 complex to the promoter, possibly by acting as a recruitment platform for the ubiquitination machinery to bind. To determine whether the Paf1 complex is indeed required for Rad6-Bre1 recruitment to promoters, ChIP experiments using TAP-tagged Rad6 were performed in a mutant missing the Rtf1 subunit of the Paf1 complex. As previously shown, Bre1 is required for the recruitment of Rad6 to promoter (Fig. 4B). However, Rad6 localization at the promoter is not affected in the absence of the Rtf1 subunit of the Paf1 complex (Fig. 4B). Thus, the mechanism by which the Paf1 complex affects monoubiquitination of histone H2B and the association of Rad6 with COMPASS and Pol II does not affect recruitment of the Rad6-Bre1 complex to the promoter region. Our results suggest that the Paf1 complex promotes ubiquitination of histone H2B by affecting the activity of the Rad6-Bre1 complex and regulating its interaction with initiating RNA polymerase II. This suggests a cooperative cross-talk mechanism between the ubiquitination and methylation machinery at the promoter regions of active genes. Following formation of the preinitiation complex, the Paf1 complex is recruited to the promoters and is required for the functional activation and ubiquitination of histone H2B by the Rad6-Bre1 complex (Fig. 5). The Paf1 complex is also involved in the subsequent association of RNA polymerase II with COMPASS (Fig. 5C). The methylation of lysines 4 and 36 of histone H3 by Set1 and Set2 has recently been linked to transcriptional elongation, and the Paf1 complex also seems to play a role in this process (17Krogan N.J. Dover J. Wood A. Schneider J. Heidt J. Boateng M.A. Dean K. Ryan O.W. Golshani A. Johnston M. Greenblatt J.F. Shilatifard A. Mol. Cell. 2003; 11: 721-729Abstract Full Text Full Text PDF PubMed Scopus (570) Google Scholar, 18Ng H.H. Robert F. Young R.A. Struhl K. Mol. Cell. 2003; 11: 709-719Abstract Full Text Full Text PDF PubMed Scopus (857) Google Scholar, 29Bernstein B.E. Humphrey E.L. Erlich R.L. Schneider R. Bouman P. Liu J.S. Kouzarides T. Schreiber S.L. Proc. Natl. Acad. Sci. U. S. A. 2002; 99: 8695-8700Crossref PubMed Scopus (601) Google Scholar, 30Santos-Rosa H. Schneider R. Bannister A.J. Sherriff J. Bernstein B.E. Emre N.C. Schreiber S.L. Mellor J. Kouzarides T. Nature. 2002; 419: 407-411Crossref PubMed Scopus (1608) Google Scholar, 32Krogan N.J. Kim M. Tong A. Golshani A. Cagney G. Canadien V. Richards D.P. Beattie B.K. Emili A. Boone C. Shilatifard A. Buratowski S. Greenblatt J. Mol. Cell. Biol. 2003; 23: 4207-4218Crossref PubMed Scopus (519) Google Scholar, 33Schaft D. Roguev A. Kotovic K.M. Shevchenko A. Sarov M. Shevchenko A. K.M. Stewart A.F. 2003; PubMed Scopus Google Scholar). Thus, the Paf1 complex plays a role in several covalent histone modifications at the promoters and the body of the each of which has in transcriptional We are to Struhl for and for We also Krogan in the Greenblatt for in the ChIP via
Ubiquitin-dependent proteolysis by the proteasome plays an essential role in a number of key biological processes, including cell cycle progression, transcription, and signal transduction (for review, see Peters et al. 1998). In many cases the target protein is first marked for degradation or processing by phosphorylation. The phosphorylated protein is then recognized and ubiquitinated in a process that requires three proteins (Hershko et al. 1983; for review, see Hershko 1998; Peters et al. 1998). Ubiquitin is first attached to a ubiquitin-activating enzyme (E1) in an ATP-dependent reaction to form a highenergy thiolester bond. The ubiquitin is then transferred from the E1 protein to an E2 ubiquitin-conjugating enzyme, which functions in conjunction with an E3 protein to link ubiquitin to lysine residues in the targeted protein. A specific lysine residue in the conjugated ubiquitin is then attached to a second ubiquitin, and reiteration of this process results in the assembly of a polyubiquitin chain. The polyubiquitinated protein can then be recognized by the 26S proteasome and degraded or processed.
In rural communities of Guerrero, internet connectivity is limited by poor wireless infrastructure. Technologies like Ubiquiti enhance coverage and support education. This project models a rural infrastructure using Ubiquiti and Cisco models, aiming to provide an accessible service. It addresses communication issues, promoting educational equity and global skills for students, making them competitive. Resources were analyzed, selecting optimal technologies, and a replicable connectivity diagram based on Cisco was designed. Connectivity with Ubiquiti benefits marginalized regions, improving online education and fostering educational, technological, and social development in underserved communities.
This study aims to implement and evaluate the performance of an inter-building wireless backhaul network using the Ubiquiti LiteBeam 5AC Gen2 integrated with a MikroTik router as a solution for internet distribution without the need to subscribe to an additional ISP service. The study is motivated by the increasing demand for high-speed and reliable network connectivity between buildings, while wired network implementations are often limited in terms of cost and installation flexibility. The research adopts an experimental approach with descriptive quantitative analysis through direct measurement of Quality of Service (QoS) parameters, including throughput, delay, and jitter. Testing was conducted under three scenarios: a direct wireless backhaul link, a WiFi network on the first floor, and a WiFi network on the second floor of the building. The results show that the wireless backhaul provides an average throughput of 87.95 Mbps for download and 48.60 Mbps for upload, with a delay of 1.19 ms. On the access network side, the achieved throughput remains sufficient for user needs, although delay and jitter increase as the number of connected devices and traffic load grows. This study concludes that the implementation of an IEEE 802.11ac-based wireless backhaul using Ubiquiti LiteBeam 5AC Gen2 and MikroTik is effective as a medium-scale inter-building connectivity solution, delivering performance that meets typical daily internet service requirements.
The ubiquitin system is complex, multifaceted, and is crucial for the modulation of a vast number of cellular processes. Ubiquitination is tightly regulated at different levels by a range of enzymes including E1s, E2s, and E3s, and an array of DUBs. The UPS directs protein degradation through the proteasome, and regulates a wide array of cellular processes including transcription and epigenetic factors as well as key oncoproteins. Ubiquitination is key to the dynamic regulation of programmed cell death. Notably, the TNF signaling pathway is controlled by competing ubiquitin conjugation and deubiquitination, which governs both proteasomal degradation and signaling complex formation. In the inflammatory response, ubiquitination is capable of both activating and dampening inflammasome activation through the control of either protein stability, complex formation, or, in some cases, directly affecting receptor activity. In this review, we discuss the enzymes and targets in the ubiquitin system that regulate fundamental cellular processes regulating cell death, and inflammation, as well as disease consequences resulting from their dysregulation. Finally, we highlight several pre-clinical and clinical compounds that regulate ubiquitin system enzymes, with the aim of restoring homeostasis and ameliorating diseases.
Histone post-transcriptional modifications play essential roles in regulation of all DNA related processes. Among them, histone ubiquitination has been discovered for more than three decades. However, its functions are still less well understood than other histone modifications such as methylation and acetylation. In this review, we will summarize our current understanding of histone ubiquitination and deubiquitination. In particular, we will focus on how they are regulated by histone ubiquitin ligases and deubiquitinating enzymes. We will then discuss the roles of histone ubiquitination in transcription and DNA damage response and the crosstalk between histone ubiquitination and other histone modifications. Finally, we will review the important roles of histone ubiquitination in stem cell biology and cancer.
The rapid development of information and communication technology has increased the need for reliable, secure, and efficient computer networks, especially for companies and organizations, including local offices. In this context, stable connectivity and optimal network management significantly impact the company's smooth operation, especially when facing increasingly growing digital transformation challenges. Companies, including the Ubiquiti device, need technology solutions to ensure smooth operations and efficiency. This study aims to provide practical solutions for companies overcoming local office network management problems. This study seeks to optimize network management in companies using Ubiquiti devices, conduct an in-depth analysis of existing network conditions, and design appropriate solutions. This study uses a descriptive qualitative approach, which aims to understand the phenomena that occur in the field in more depth. This approach was chosen because of its ability to explore the meaning, processes, and dynamics that occur in a particular social context. To ensure the successful implementation of the proposed solution, here are some suggestions that PT. Rizki Inti Madani can consider. Human Resources Training and Development, considering the importance of a deep understanding of devices and centralized network management, it is highly recommended that IT technicians and staff involved in network management be given further training on using Ubiquiti and Cisco devices. This training should also include understanding VLAN configuration, QoS, and efficient network troubleshooting and monitoring techniques. Gradual Transition: to reduce the risk of operational disruption, it is recommended that the implementation be carried out in stages. The devices will first be tested in a laboratory environment to ensure the configuration is running as expected. After successful testing, the devices can be installed gradually throughout the office to minimize downtime and operational disruption.
The background of this research is the increasing demand and competition in the wireless network device industry. Ubiquiti, as one of the major players in the industry, needs to understand how brand image, brand awareness, and promotional activities influence consumers' purchasing decisions. This study aims to analyze the influence of brand image, brand awareness, and promotion variables on purchasing decisions on Ubiquiti brand IT network products. Data were collected from 97 respondents, and analysis was conducted on the demographics of respondents, product use, and the influence of independent variables on purchasing decisions. Research analysis method using SPSS. The results showed that the majority of product users were men aged 25-29 years who worked as employees and used the product for less than 1 year. Statistical analysis revealed that the three variables of brand image, brand awareness, and promotion significantly influence purchasing decisions. Brand awareness has been shown to have the most influence, followed by brand image and promotion. This finding is reinforced by brand awareness's largest adjusted R square value (96.8%). The main conclusion is that brand awareness has the most significant influence on consumer purchasing decisions Ubiquiti brand IT network products, followed by brand image and promotion. The implication of this study is the importance of companies in designing effective marketing and branding strategies to improve consumer purchasing decisions. The research provides valuable insights for Ubiquiti and other companies in the wireless networking industry to better understand the factors driving consumer buying behavior, so they can develop targeted marketing strategies and increase their market share.
Assess the impact of heterogeneity among established sepsis criteria (Sepsis-1, Sepsis-3, Centers for Disease Control and Prevention Adult Sepsis Event, and Centers for Medicare and Medicaid severe sepsis core measure 1) through the comparison of corresponding sepsis cohorts. Retrospective analysis of data extracted from electronic health record. Single, tertiary-care center in St. Louis, MO. Adult, nonsurgical inpatients admitted between January 1, 2012, and January 6, 2018. None. In the electronic health record data, 286,759 encounters met inclusion criteria across the study period. Application of established sepsis criteria yielded cohorts varying in prevalence: Centers for Disease Control and Prevention Adult Sepsis Event (4.4%), Centers for Medicare and Medicaid severe sepsis core measure 1 (4.8%), International Classification of Disease code (7.2%), Sepsis-3 (7.5%), and Sepsis-1 (11.3%). Between the two modern established criteria, Sepsis-3 (n = 21,550) and Centers for Disease Control and Prevention Adult Sepsis Event (n = 12,494), the size of the overlap was 7,763. The sepsis cohorts also varied in time from admission to sepsis onset (hr): Sepsis-1 (2.9), Sepsis-3 (4.1), Centers for Disease Control and Prevention Adult Sepsis Event (4.6), and Centers for Medicare and Medicaid severe sepsis core measure 1 (7.6); sepsis discharge International Classification of Disease code rate: Sepsis-1 (37.4%), Sepsis-3 (40.1%), Centers for Medicare and Medicaid severe sepsis core measure 1 (48.5%), and Centers for Disease Control and Prevention Adult Sepsis Event (54.5%); and inhospital mortality rate: Sepsis-1 (13.6%), Sepsis-3 (18.8%), International Classification of Disease code (20.4%), Centers for Medicare and Medicaid severe sepsis core measure 1 (22.5%), and Centers for Disease Control and Prevention Adult Sepsis Event (24.1%). The application of commonly used sepsis definitions on a single population produced sepsis cohorts with low agreement, significantly different baseline demographics, and clinical outcomes.
We prove that the consistently oriented double ray is ubiquitous if and only if it is ubiquitous restricted to the class of one-ended digraphs. Additionally, we prove the same equivalence for the disjoint union of a consistently forward and a consistently backward oriented ray. Furthermore, we discuss the connection between these two ubiquity problems.
Let R be a d-dimensional Cohen-Macaulay complete local ring with infinite residue field k. The dominant index $\operatorname{dx}(R)$ is by definition the least number of extensions necessary to build k in the singularity category $\operatorname{D^{sg}}$ out of each nonzero object, up to finite direct sums, direct summands and shifts. The local ring R is called uniformly dominant if $\operatorname{dx}(R)$ is finite. In this paper, we prove that R is uniformly dominant with $\operatorname{dx}(R)\le6d+5$ if R has codimension 2 and is not a complete intersection. Also, we show that R is uniformly dominant with $\operatorname{dx}(R)\le d+1$ if R is Burch, and with $\operatorname{dx}(R)\le d$ if R is either a quasi-fiber product ring, or has multiplicity at most 5 and is not Gorenstein. A result on hypersurfaces by Ballard, Favero and Katzarkov is recovered, and results on Burch rings and quasi-fiber product rings by Takahashi are refined.
Complex contagions describe systems where the probability or rate of contagious transmission is a nonlinear function of the exposure to contagious agents. These models were first studied theoretically but have since been used to capture effects such as nonconformism, social reinforcement or peer pressure in empirical data. However, recent studies have shown that local correlations (e.g., group structure or temporal burstiness) and heterogeneity (e.g., diversity of parameters or covariates) can give the illusion of nonlinear effects even when the dynamics is actually linear. We briefly review these studies to inform a new model and explanation for these effective models of complex contagions. We find global threshold dynamics and superlinear complex contagions even in populations where agents are distributed across social groups described solely by linear or even sublinear contagions. This effect can be understood as a manifestation of Simpson's paradox. Incidence data from heterogeneous groups can look superlinear once averaged over all groups, since the sampling of groups represented at high incidence is biased towards those with stronger local transmission. We then define what we
Complex organic molecules (COMs) in starless cores provide critical insights into the early stages of star formation and prebiotic chemistry. We present a chemical survey of 16 starless cores (including five prestellar cores) in the Orion A and B molecular clouds, targeting CH3OH, N2H+, CCS, and c-C3HD, using the Atacama Compact Array (ACA) and the Yebes 40-m telescope. CH3OH was detected toward all targets, confirming its ubiquity in starless cores, consistent with previous surveys in Taurus and Perseus. ACA imaging shows that CH3OH, CCS, and c-C3HD generally trace the outer layers of the dense cores outlined by N2H+, each exhibiting distinct spatial distributions. Meanwhile, Comparison with Yebes data reveals an extended, flattened CH3OH component. CCS and c-C3HD tend to be detected or non-detected together across cores, while cores near dust-rich regions on a large scale often lack both, suggesting environmental influences linked to the interstellar radiation field. Within individual cores, CCS typically resides in an outer layer relative to c-C3HD. Our findings underscore the importance of high-resolution studies for understanding the origins and spatial differentiation of COMs
We demonstrate that final-state uncertainty is ubiquitous in multistable systems of coupled neuronal maps, meaning that predicting whether one such system will eventually be chaotic or nonchaotic is often nearly impossible. We propose a "chance synchronization" mechanism that governs the emergence of unpredictability in neuron systems and support it by using basin classification, uncertainty exponent, and basin entropy techniques to analyze five simple discrete-time systems, each consisting of a different neuron model. Our results illustrate that uncertainty in neuron systems is not just a product of noise or high-dimensional complexity; it is also a fundamental property of low-dimensional, deterministic models, which has profound implications for understanding brain function, modeling cognition, and interpreting unpredictability in general multistable systems.
The Neural Tangent Kernel theory theoretically guarantees the existence of global minima of the cost functional in the neighborhood of an arbitrary random initialized parameters in wide artificial neural networks. In this paper, we show that the ubiquity of the global minima directly corresponds to the typicality of pure thermal states in isolated quantum systems by identifying a common underlying mechanism characterized by the restriction to a few observables and the role of a Wishart-type matrix. Moreover, we demonstrate that the increase in distinguishability of the reduced density matrices of typical pure states with subsystem size corresponds to the double descent phenomenon observed by varying the width of layers in finite-width artificial neural networks. Thereby, the threshold for the reduced state become thermal is determined by essentially the same condition as the fitting threshold. In this manner, we reveal a structural correspondence between thermalization in isolated quantum systems and wide neural network.
We present high-resolution optical emission spectroscopy observations of the ultra hot Jupiters (UHJs) TOI-1431 b and TOI-1518 b using the PEPSI spectrograph on the LBT. We detect emission lines from Fe I with a significance of 5.68 $σ$ and 7.68 $σ$ for TOI 1431 b and TOI-1518 b, respectively. We also tentatively detect Cr I emission from TOI-1431 b at $4.32σ$. For TOI-1518 b, we tentatively detect Ni I, Fe II, and Mg I at significance levels ranging from $3-4σ$. Detection of emission lines indicates that both planets possess temperature inversions in their atmospheres, providing further evidence of the ubiquity of stratospheres among UHJs. By analyzing the population of hot Jupiters, we compare models that predict the distribution of planets in the temperature-gravity space, and find a recent global circulation model suite from Roth et al. (2024) provides a reasonable match to the observed onset of inversions at $T_{\mathrm{eq}}\sim2000$ K. The ubiquity of strong Fe I emission lines among UHJs, together with the paucity of detections of TiO, suggest that atomic iron is the dominant optical opacity source in their atmospheres and can be responsible for the inversions.
Models and observations have demonstrated that Twisted Flux Ropes (TFRs) play a significant role in the structure and eruptive dynamics of active regions. Their role in the dynamics of the quiet Sun atmosphere on has remained elusive, their fundamental relevance emerging mainly from theoretical models (Amari et al. 2015), showing that they form and erupt as a result of flux cancellation. Here HINODE high-resolution photospheric vector magnetic field measurements are integrated with advanced environment reconstruction models: TFRs develop on various scales and are associated with the appearance of mesospots. The developing TFRs contain sufficient free magnetic energy to match the requirements of the recently observed "campfires" discovered by Solar Orbiter in the quiet Sun. The free magnetic energy is found to be large enough to trigger eruptions while the magnetic twist large enough to trigger confined eruptions, heating the atmosphere. TFRs are also connected to larger scale magnetic fields such as supergranulation loops, allowing the generation of Alfvén waves at the top of the chromosphere that can propagate along them. High-resolution magnetohydrodynamic simulations, incorporat
A digraph $H$ is called ubiquitous if every digraph that contains arbitrarily many vertex-disjoint copies of $H$ also contains infinitely many vertex-disjoint copies of $H$. We study oriented double rays, that is, digraphs $H$ whose underlying undirected graphs are double rays. Calling a vertex of an oriented double ray a turn if it has in-degree or out-degree 2, we prove that an oriented double ray with at least one turn is ubiquitous if and only if it has a (finite) odd number of turns. It remains an open problem to determine whether the consistently oriented double ray is ubiquitous.