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Karen E Hayes, Jamie A Barr, Mingyi Xie, Joan A Steitz, Ivan Martinez
Cellular quiescence (also known as G0 arrest) is characterized by reduced DNA replication, increased autophagy, and increased expression of cyclin-dependent kinase p27Kip1 . Quiescence is essential for wound healing, organ regeneration, and preventing neoplasia. Previous findings indicate that microRNAs (miRNAs) play an important role in regulating cellular quiescence. Our recent publication demonstrated the existence of an alternative miRNA biogenesis pathway in primary human foreskin fibroblast (HFF) cells during quiescence...
February 5, 2018: Bio-protocol
Anne-Sophie Gribling-Burrer, Gilbert Eriani, Christine Allmang
Several selenoprotein mRNAs undergo 5' cap maturation events whereby their classical monomethylated m(7)G cap becomes trimethylated (m3(2,2,7)G) by the trimethylguanosine synthase 1 (Tgs1). Here, we describe immunoprecipitation methods for the detection of endogenous m3(2,2,7)G-capped selenoprotein mRNAs from total cell extracts or after polysome fractionation of cytoplasmic extracts. We have also developed a method for the in vitro cap hypermethylation of selenoprotein mRNA transcripts using purified Tgs1 enzyme...
2018: Methods in Molecular Biology
Priyanka Kumari, Kriti Sikri, Kohinoor Kaur, Umesh Datta Gupta, Jaya Sivaswami Tyagi
DevR/DosR is a key mediator of 'dormancy' adaptation in Mycobacterium tuberculosis in response to gaseous stresses such as hypoxia that inhibit aerobic mode of respiration. In the present study, a temporal analysis over a 1 year period has revealed robust expression of representative DevR regulon genes devR, hspX and tgs1, during long-term 'dormancy' adaptation to hypoxia. Notably, a predominant proportion of long-term hypoxia-adapted bacteria were characterized by their inability to grow on solid media, accumulation of triacylglycerols and recovery of growth in liquid media...
September 2017: Tuberculosis
Jinpeng Gao, James G Wallis, Jeremy B Jewell, John Browse
Chilling stress is a major factor limiting plant development and crop productivity. Because the plant response to chilling is so complex, we are far from understanding the genes important in the response to chilling. To identify new genes important in chilling tolerance, we conducted a novel mutant screen, combining a confirmed SALK T-DNA insertion collection with traditional forward genetics. We screened a pool of more than 3700 confirmed homozygous SALK T-DNA insertion lines for visible defects under prolonged growth at 5°C...
July 2017: Plant Physiology
Anne-Sophie Gribling-Burrer, Michael Leichter, Laurence Wurth, Alexandra Huttin, Florence Schlotter, Nathalie Troffer-Charlier, Vincent Cura, Martine Barkats, Jean Cavarelli, Séverine Massenet, Christine Allmang
Selenoprotein synthesis requires the co-translational recoding of a UGASec codon. This process involves an RNA structural element, called Selenocysteine Insertion Sequence (SECIS) and the SECIS binding protein 2 (SBP2). Several selenoprotein mRNAs undergo unusual cap hypermethylation by the trimethylguanosine synthase 1 (Tgs1), which is recruited by the ubiquitous Survival of MotoNeurons (SMN) protein. SMN, the protein involved in spinal muscular atrophy, is part of a chaperone complex that collaborates with the methylosome for RNP assembly...
May 19, 2017: Nucleic Acids Research
Prasad H Babar, Vishakha Dey, Praveen Jaiswar, Swati Patankar
Many Plasmodium falciparum proteins do not share homology with, and are generally longer than their respective orthologs. This, to some extent, can be attributed to insertions. Here, we studied a P. falciparum RNA hypermethylase, trimethylguanosine synthase (PfTGS1) that harbors a 76 amino acid insertion in its methyltransferase domain. Bioinformatics analysis revealed that this insertion was present in TGS1 orthologs from other Plasmodium species as well. Interestingly, a classical nuclear localization signal (NLS) was predicted in the insertions of primate parasite TGS1 proteins...
November 2016: Molecular and Biochemical Parasitology
Albertus Viljoen, Mickael Blaise, Chantal de Chastellier, Laurent Kremer
Slow growing pathogenic mycobacteria utilize host-derived lipids and accumulate large amounts of triacylglycerol (TAG) in the form of intracytoplasmic lipid inclusions (ILI), serving as a source of carbon and energy during prolonged infection. Mycobacterium abscessus is an emerging and rapidly growing species capable to induce severe and chronic pulmonary infections. However, whether M. abscessus, like Mycobacterium tuberculosis, possesses the machinery to acquire and store host lipids, remains unaddressed...
November 2016: Molecular Microbiology
Rebecca M Borg, Benji Fenech Salerno, Neville Vassallo, Rémy Bordonne, Ruben J Cauchi
The neuromuscular disorder, spinal muscular atrophy (SMA), results from insufficient levels of the survival motor neuron (SMN) protein. Together with Gemins 2-8 and Unrip, SMN forms the large macromolecular SMN-Gemins complex, which is known to be indispensable for chaperoning the assembly of spliceosomal small nuclear ribonucleoproteins (snRNPs). It remains unclear whether disruption of this function is responsible for the selective neuromuscular degeneration in SMA. In the present study, we first show that loss of wmd, the Drosophila Unrip orthologue, has a negative impact on the motor system...
October 2016: Neurobiology of Disease
Su-Dong Liu, Jin Su, Shi-Meng Zhang, Hai-Ping Dong, Hui Wang, Wei Luo, Qian Wen, Jian-Chun He, Xiao-Fan Yang, Li Ma
New vaccines are needed to combat Mycobacterium tuberculosis (MTB) infections. The currently employed Bacillus Calmette-Guérin vaccine is becoming ineffective, due in part to the emergence of multidrug-resistant tuberculosis (MDR-TB) strains and the reduced immune capacity in cases of HIV coinfection. CD8(+) T cells play an important role in the protective immunity against MTB infections, and the identification of immunogenic CD8(+) T cell epitopes specific for MTB is essential for the design of peptide-based vaccines...
September 2016: Journal of Cellular and Molecular Medicine
Shivangi Rastogi, Pooja Agarwal, Manju Y Krishnan
During its persistence in the infected host, Mycobacterium tuberculosis (Mtb) accumulates host-derived fatty acids in intracytoplasmic lipid inclusions as triacylglycerols which serve primarily as carbon and energy reserves. The Mtb genome codes for more than 15 triacylglycerol synthases, 24 lipase/esterases, and seven cutinase-like proteins. Hence, we looked at the expression of the corresponding genes in intracellular bacilli persisting amidst the host triacylglycerols. We used the Mtb infected murine adipocyte model to ensure persistence and transcripts were quantified using real-time reverse transcriptase polymerase chain reaction...
March 2016: International Journal of Mycobacteriology
Weichao Huang, Carolina Río Bártulos, Peter G Kroth
Diatoms are unicellular photoautotrophic algae, which can be found in any aquatic habitat. The main storage carbohydrate of diatoms is chrysolaminarin, a nonlinear β-glucan, consisting of a linear 1,3-β-chain with 1,6-β-branches, which is stored in cytoplasmic vacuoles. The metabolic pathways of chrysolaminarin synthesis in diatoms are poorly investigated, therefore we studied two potential 1,6-β-transglycosylases (TGS) of the diatom Phaeodactylum tricornutum which are similar to yeast Kre6 proteins and which potentially are involved in the branching of 1,3-β-glucan chains by adding d-glucose as 1,6-side chains...
July 2016: Journal of Eukaryotic Microbiology
Fan Yan, Shaokang Di, Ryoji Takahashi
The R gene of soybean, presumably encoding a MYB transcription factor, controls seed coat color. The gene consists of multiple alleles, R (black), r-m (black spots and (or) concentric streaks on brown seed), and r (brown seed). This study was conducted to determine the structure of the MYB transcription factor gene in a near-isogenic line (NIL) having r-m allele. PCR amplification of a fragment of the candidate gene Glyma.09G235100 generated a fragment of about 1 kb in the soybean cultivar Clark, whereas a fragment of about 14 kb in addition to fragments of 1 and 1...
August 2015: Genome Génome / Conseil National de Recherches Canada
Takashi Nakatsuka, Misa Saito, Eri Yamada, Kohei Fujita, Noriko Yamagishi, Nobuyuki Yoshikawa, Masahiro Nishihara
BACKGROUND: Generally, double-flowered varieties are more attractive than single-flowered varieties in ornamental plants. Japanese gentian is one of the most popular floricultural plants in Japan, and it is desirable to breed elite double-flowered cultivars. In this study, we attempted to characterize a doubled-flower mutant of Japanese gentian. To identify the gene that causes the double-flowered phenotype in Japanese gentian, we isolated and characterized MADS-box genes. RESULTS: Fourteen MADS-box genes were isolated, and two of them were C-class MADS-box genes (GsAG1 and GsAG2)...
2015: BMC Plant Biology
Kum-Loong Boon, Michael David Pearson, Martin Koš
Trimethylguanosine Synthase catalyses transfer of two methyl groups to the m(7)G cap of RNA polymerase II transcribed snRNAs, snoRNAs, and telomerase RNA TLC1 to form a 2,2,7-trimethylguanosine cap. While in vitro studies indicate that Tgs1 functions as a monomer and the dimethylation of m(7)G caps is not a processive reaction, partially methylated sn(o)RNAs are typically not detected in living cells. Here we show that both yeast and human Tgs1p possess a conserved self-association property located at the N-terminus...
2015: Scientific Reports
Zhicheng R Qiu, Beate Schwer, Stewart Shuman
The trimethylguanosine (TMG) caps of small nuclear (sn) RNAs are synthesized by the enzyme Tgs1 via sequential methyl additions to the N2 atom of the m(7)G cap. Whereas TMG caps are inessential for Saccharomyces cerevisiae vegetative growth at 25° to 37°, tgs1∆ cells that lack TMG caps fail to thrive at 18°. The cold-sensitive defect correlates with ectopic stoichiometric association of nuclear cap-binding complex (CBC) with the residual m(7)G cap of the U1 snRNA and is suppressed fully by Cbc2 mutations that weaken cap binding...
July 2015: G3: Genes—Genomes—Genetics
Tatsufumi Murakami, Kazunori Sango, Kazuhiko Watabe, Naoko Niimi, Shizuka Takaku, Zhenghua Li, Ken-ichi Yamamura, Yoshihide Sunada
Familial amyloidotic polyneuropathy (FAP) is one of the transthyretin (TTR) amyloidoses characterized by extracellular amyloid deposits and peripheral nerve involvement. Recently, we found significant expression of the TTR gene in Schwann cells of the peripheral nervous system. We hypothesized that local expression of variant TTR in Schwann cells may contribute to neurodegeneration in FAP. Schwann cells derived from the dorsal root ganglia (DRG) of transgenic mice expressing variant human TTR in a mouse null background were cultured long term to obtain spontaneously immortalized cell lines...
July 2015: Journal of Neurochemistry
Laurence Wurth, Anne-Sophie Gribling-Burrer, Céline Verheggen, Michael Leichter, Akiko Takeuchi, Stéphanie Baudrey, Franck Martin, Alain Krol, Edouard Bertrand, Christine Allmang
Mammalian mRNAs are generated by complex and coordinated biogenesis pathways and acquire 5'-end m(7)G caps that play fundamental roles in processing and translation. Here we show that several selenoprotein mRNAs are not recognized efficiently by translation initiation factor eIF4E because they bear a hypermethylated cap. This cap modification is acquired via a 5'-end maturation pathway similar to that of the small nucle(ol)ar RNAs (sn- and snoRNAs). Our findings also establish that the trimethylguanosine synthase 1 (Tgs1) interacts with selenoprotein mRNAs for cap hypermethylation and that assembly chaperones and core proteins devoted to sn- and snoRNP maturation contribute to recruiting Tgs1 to selenoprotein mRNPs...
July 2014: Nucleic Acids Research
Y Ramayo-Caldas, M R S Fortes, N J Hudson, L R Porto-Neto, S Bolormaa, W Barendse, M Kelly, S S Moore, M E Goddard, S A Lehnert, A Reverter
High intramuscular fat (IMF) awards price premiums to beef producers and is associated with meat quality and flavor. Studying gene interactions and pathways that affect IMF might unveil causative physiological mechanisms and inform genomic selection, leading to increased accuracy of predictions of breeding value. To study gene interactions and pathways, a gene network was derived from genetic markers associated with direct measures of IMF, other fat phenotypes, feedlot performance, and a number of meat quality traits relating to body conformation, development, and metabolism that might be plausibly expected to interact with IMF biology...
July 2014: Journal of Animal Science
Beate Schwer, Stewart Shuman
Yhc1 and U1C are homologous essential subunits of the yeast and human U1 snRNP, respectively, that are implicated in the establishment and stability of the complex of U1 bound to the pre-mRNA 5' splice site (5'SS). Here, we conducted a mutational analysis of Yhc1, guided by the U1C NMR structure and low-resolution crystal structure of human U1 snRNP. The N-terminal 170-amino acid segment of the 231-amino acid Yhc1 polypeptide sufficed for vegetative growth. Although changing the zinc-binding residue Cys6 to alanine was lethal, alanines at zinc-binding residues Cys9, His24 and His30 were not...
April 2014: Nucleic Acids Research
Nidhi Kapoor, Santosh Pawar, Tatiana D Sirakova, Chirajyoti Deb, William L Warren, Pappachan E Kolattukudy
Tuberculosis (TB) is responsible for death of nearly two million people in the world annually. Upon infection, Mycobacterium tuberculosis (Mtb) causes formation of granuloma where the pathogen goes into dormant state and can live for decades before resuscitation to develop active disease when the immune system of the host is weakened and/or suppressed. In an attempt to better understand host-pathogen interactions, several groups have been developing in vitro models of human tuberculosis granuloma. However, to date, an in vitro granuloma model in which Mtb goes into dormancy and can subsequently resuscitate under conditions that mimic weakening of the immune system has not been reported...
2013: PloS One
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