Histone 2B monoubiquitination complex integrates transcript elongation with RNA processing at circadian clock and flowering regulators

Altres ajuts: CERCA Programme/Generalitat de CatalunyaHISTONE MONOUBIQUITINATION1 (HUB1) and its paralog HUB2 act in a conserved heterotetrameric complex in the chromatin-mediated transcriptional modulation of developmental programs, such as flowering time, dormancy, and the circadian clock. The KHD1 and SPEN3 proteins were identified as interactors of the HUB1 and HUB2 proteins with in vitro RNA-binding activity. Mutants in SPEN3 and KHD1 had reduced rosette and leaf areas. Strikingly, in spen3 mutants, the flowering time was slightly, but significantly, delayed, as opposed to the early flowering time in the hub1-4 mutant. The mutant phenotypes in biomass and flowering time suggested a deregulation of their respective regulatory genes CIRCADIAN CLOCK-ASSOCIATED1 (CCA1) and FLOWERING LOCUS C (FLC) that are known targets of the HUB1-mediated histone H2B monoubiquitination (H2Bub). Indeed, in the spen3-1 and hub1-4 mutants, the circadian clock period was shortened as observed by luciferase reporter assays, the levels of the CCA1α and CCA1β splice forms were altered, and the CCA1 expression and H2Bub levels were reduced. In the spen3-1 mutant, the delay in flowering time was correlated with an enhanced FLC expression, possibly due to an increased distal versus proximal ratio of its antisense COOLAIR transcript. Together with transcriptomic and double-mutant analyses, our data revealed that the HUB1 interaction with SPEN3 links H2Bub during transcript elongation with pre-mRNA processing at CCA1. Furthermore, the presence of an intact HUB1 at the FLC is required for SPEN3 function in the formation of the FLC-derived antisense COOLAIR transcripts

Impaired CK1 Delta Activity Attenuates SV40-Induced Cellular Transformation In Vitro and Mouse Mammary Carcinogenesis In Vivo

Simian virus 40 (SV40) is a powerful tool to study cellular transformation in vitro, as well as tumor development and progression in vivo. Various cellular kinases, among them members of the CK1 family, play an important role in modulating the transforming activity of SV40, including the transforming activity of T-Ag, the major transforming protein of SV40, itself. Here we characterized the effects of mutant CK1δ variants with impaired kinase activity on SV40-induced cell transformation in vitro, and on SV40-induced mammary carcinogenesis in vivo in a transgenic/bi-transgenic mouse model. CK1δ mutants exhibited a reduced kinase activity compared to wtCK1δ in in vitro kinase assays. Molecular modeling studies suggested that mutation N172D, located within the substrate binding region, is mainly responsible for impaired mutCK1δ activity. When stably over-expressed in maximal transformed SV-52 cells, CK1δ mutants induced reversion to a minimal transformed phenotype by dominant-negative interference with endogenous wtCK1δ. To characterize the effects of CK1δ on SV40-induced mammary carcinogenesis, we generated transgenic mice expressing mutant CK1δ under the control of the whey acidic protein (WAP) gene promoter, and crossed them with SV40 transgenic WAP-T-antigen (WAP-T) mice. Both WAP-T mice as well as WAP-mutCK1δ/WAP-T bi-transgenic mice developed breast cancer. However, tumor incidence was lower and life span was significantly longer in WAP-mutCK1δ/WAP-T bi-transgenic animals. The reduced CK1δ activity did not affect early lesion formation during tumorigenesis, suggesting that impaired CK1δ activity reduces the probability for outgrowth of in situ carcinomas to invasive carcinomas. The different tumorigenic potential of SV40 in WAP-T and WAP-mutCK1δ/WAP-T tumors was also reflected by a significantly different expression of various genes known to be involved in tumor progression, specifically of those involved in wnt-signaling and DNA repair. Our data show that inactivating mutations in CK1δ impair SV40-induced cellular transformation in vitro and mouse mammary carcinogenesis in vivo

İyileştirilmiş paylaşımlı küme kullanımı için melez iş çizelgelemesi

Thesis (M.A.)--Özyeğin University, Graduate School of Sciences and Engineering, Department of Computer Science, January 2014.Bu tezde, bilgisayar küme yapıları üzerinde hibrid iş yüklerinin birlikte işlenmesiyle ilgili modeller, sorunlar ve performans kazanımları incelenmektedir. Desteklenen kümeleme teknolojileri arasında MPI, Hadoop-MapReduce ve NoSQL sistemleri bulunmaktadır. Önerilen programlayıcı modeli işletim sistemi seviyesindeki arakatman yazılımların üzerinde ve onları destekleyici niteliktedir. Tezde ilk olarak, MPI,Hadoop ve NoSQL işlerini bir arada programlayabildiğimizi göstermekteyiz.İkinci olarak, farklı özelliklere sahip (CPU vs. Girdi/Çıktı yoğunluklu) işlerin, aynı özelliklere sahip işlere göre (2 adet veya daha fazla CPU yoğunluklu) beraber daha iyi programlanabildiği bulgusu paylaşılmaktadır. Son olarak, bu bulgunun ışığında yeni bir greedy sort-merge programlayıcısı tasarımı anlatılmaktadır. İş tamamlama sürelerinde %37 zamansal kazanım gösterilmektedir, ancak %50 kazanımlar da (2x hızlanma) teorik olarak mümkündür. Bu zamansal kazanımlar kuyrukta yeterince yük olduğu takdirde kümenin kullanım kapasitesini de arttırıcı nitelikte olacaktır. Tezin sonunda, hibrid iş programlama ile sağlanabilecek potansiyel güç-enerji kazanımları da tartışılmaktadır.In this thesis, We investigate the models and issues as well as performance benefits of hybrid job scheduling over shared physical clusters. Clustering technologies that are compared include MPI, Hadoop-MapReduce and NoSQL systems. Our proposed scheduling model is above the operating system and cluster-middleware level job schedulers and operating system level schedulers and it is complementary to them. First, we demonstrate that we can schedule MPI, Hadoop and NoSQL cluster-level jobs together in a controlled-fashion over the same physical cluster. Second, we find that it is better to schedule cluster jobs with different job characteristics together (CPU vs. I/O intensive) rather than two or more CPU intensive jobs. Third, we describe the design of a greedy sort-merge scheduler that uses the learning outcome of this principle. Up to 37% savings in total job completion times are demonstrated for I/O and CPU-intensive pairs of jobs, but up to 50% savings (or 2x speedup) is theoretically possible. These savings would also be proportional to the cluster utilization improvements, if there are jobs waiting in the queue. At the end of the thesis, we also discuss potential power-energy savings from hybrid job scheduling.TÜBİAK ; European Commissio

Transcript Elongation Factor TFIIS Is Involved in Arabidopsis Seed Dormancy

Grasser M, Kane CM, Merkle T, Melzer M, Emmersen J, Grasser KD. Transcript Elongation Factor TFIIS Is Involved in Arabidopsis Seed Dormancy. Journal of Molecular Biology. 2009;386(3):598-611.Transcript elongation factor TFIIS promotes efficient transcription by RNA polymerase II, since it assists in bypassing blocks during mRNA synthesis. While yeast cells lacking TFIIS are viable, inactivation of mouse TFIIS causes embryonic lethality. Here, we have identified a protein encoded in the Arabidopsis genome that displays a marked sequence similarity to TFIIS of other organisms, primarily within domains II and III in the C-terminal part of the protein. TFIIS is widely expressed in Arabidopsis, and a green fluorescent protein-TFIIS fusion protein localises specifically to the cell nucleus. When expressed in yeast cells lacking the endogenous TFIIS, Arabidopsis TFIIS partially complements the sensitivity of mutant cells to the nucleotide analog 6-azauridine, which is a typical characteristic of transcript elongation factors. We have characterised Arabidopsis lines harbouring T-DNA insertions in the coding sequence of TFIIS. Plants homozygous for T-DNA insertions are viable, and genomewide transcript profiling revealed that compared to control plants, a relatively small number of genes are differentially expressed in mutant plants. TFIIS-/- plants display essentially normal development, but they flower slightly earlier than control plants and show clearly reduced seed dormancy. Plants with RNAi-mediated knockdown of TFIIS expression also are affected in seed dormancy. Therefore, TFIIS plays a critical role in Arabidopsis seed development. (c) 2008 Elsevier Ltd. All rights reserved

Reduced expression of the DOG1 gene in Arabidopsis mutant seeds lacking the transcript elongation factor TFIIS

AbstractTFIIS is a transcript elongation factor that facilitates transcription by RNA polymerase II through blocks to elongation. Arabidopsis plants lacking TFIIS are affected in seed dormancy, which represents a block to complete germination under favourable conditions. We have comparatively profiled the transcript levels of seeds of tfIIs mutants and control plants. Among the differentially expressed genes, the DOG1 gene was identified that is a QTL for seed dormancy. The reduced expression of DOG1 in tfIIs seeds was confirmed by quantitative RT-PCR and Northern analyses, suggesting that down-regulation of DOG1 expression is involved in the seed dormancy phenotype of tfIIs mutants

ArabidopsismRNA export factor MOS11: molecular interactions and role in abiotic stress responses

� Transcription and export (TREX) is a multi-subunit complex that links synthesis, processing and export of mRNAs. It interacts with the RNA helicase UAP56 and export factors such as MOS11 and ALYs to facilitate nucleocytosolic transport of mRNAs. Plant MOS11 is a conserved, but sparsely researched RNA-binding export factor, related to yeast Tho1 and mammalian CIP29/SARNP. � Using biochemical approaches, the domains of Arabidopsis thaliana MOS11 required for interaction with UAP56 and RNA-binding were identified. Further analyses revealed marked genetic interactions between MOS11 and ALY genes. Cell fractionation in combination with transcript profiling demonstrated that MOS11 is required for export of a subset of mRNAs that are shorter and more GC-rich than MOS11-independent transcripts. � The central a-helical domain of MOS11 proved essential for physical interaction with UAP56 and for RNA-binding. MOS11 is involved in the nucleocytosolic transport of mRNAs that are upregulated under stress conditions and accordingly mos11 mutant plants turned out to be sensitive to elevated NaCl concentrations and heat stress. � Collectively, our analyses identify functional interaction domains of MOS11. In addition, the results establish that mRNA export is critically involved in the plant response to stress conditions and that MOS11 plays a prominent role at this

The UAP56-Interacting Export Factors UIEF1 and UIEF2 Function in mRNA Export

In eukaryotes, the regulated transport of mRNAs from the nucleus to the cytosol through nuclear pore complexes represents an important step in the expression of protein-coding genes. In plants, the mechanism of nucleocytosolic mRNA transport and the factors involved are poorly understood. The Arabidopsis (Arabidopsis thaliana) genome encodes two likely orthologs of UAP56-interacting factor, which acts as mRNA export factor in mammalian cells. In yeast and plant cells, both proteins interact directly with the mRNA export-related RNA helicase UAP56 and the interaction was mediated by an N-terminal UAP56-binding motif. Accordingly, the two proteins were termed UAP56-INTERACTING EXPORT FACTOR1 and 2 (UIEF1/2). Despite lacking a known RNA-binding motif, recombinant UIEF1 interacted with RNA, and the C-terminal part of UIEF1 mainly contributed to the RNA interaction. Mutation of UIEF1, UIEF2, or both in the double-mutant 2xuief caused modest growth defects. A cross between the 2xuief and 4xaly (defective in the four ALY1-4 mRNA export factors) mutants produced the sextuple mutant 4xaly 2xuief, which displayed more severe growth impairment than the 4xaly plants. Developmental defects including delayed bolting and reduced seed set were observed in the 4xaly but not the 2xuief plants. Analysis of the cellular distribution of polyadenylated mRNAs revealed more pronounced nuclear mRNA accumulation in 4xaly 2xuief than in 2xuief and 4xaly cells. In conclusion, the results indicate that UIEF1 and UIEF2 act as mRNA export factors in plants and that they cooperate with ALY1-ALY4 to mediate efficient nucleocytosolic mRNA transport

Comparison of the transcriptomes of tfIIs vs Col-0

Accession Number: GSE10663 Platform: GPL6177: Agilent-015059 Arabidopsis 3 Oligo Microarray 4x44K G2519F (Feature Number version) Organism: Arabidopsis thaliana Published on 2009-03-24 Summary: Testing the consequences of the absence of the transcript elongation factor TFIIS on the transcriptome in Arabidopsis. Keywords: Comparison of mutant and wildtype Overall Design: Four replicate RNA extractions were performed of each genotype using independent pools of plants. Samples were hybridized in two dye-swaps. tfIIs1#1 (Cy5) vs. Col-0#1 (Cy3) Col-0#2 (Cy5) vs. tfIIs1#2 (Cy3) dye-swap tfIIs1#3 (Cy5) vs. Col-0#3 (Cy3) Col-0#5 (Cy5) vs. tfIIs1#4 (Cy3) dye-swap Samples were hybridized in two dye-swaps. The data were normalized over all 4 hybridizations to obtain one, single log-ratio(Sample/Reference). The raw data files of all four of the hybridizations are attached to the Sample. Contact: Name: Wouter Van Delm Organization: Flanders Institute for Biotechnology (VIB) Deparment: Nucleomics Core Address: Herestraat 49 Box 816 Leuven Belgium Email: [email protected] Phone: +32(0)16 37 31 26 Organization: Agilent Technologies Address: Palo Alto CA 94304 USA Email: [email protected] Phone: 877-424-4536 Web-Link: www.agilent.co