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Arc is a unique immediate early gene (IEG) whose expression is induced as synapses are modified during learning. Newly-synthesized Arc mRNA is rapidly transported throughout dendrites and localizes near recently activated synapses. Arc mRNA levels are regulated by rapid degradation, which is accelerated by synaptic activity in a translation-dependent process. One possible mechanism is nonsense-mediated mRNA decay (NMD), which depends on the presence of a splice junction in the 3'UTR. Here, we test this hypothesis using transgenic mice that express EGFP-Arc. Because the transgene was constructed from Arc cDNA, it lacks intron structures in the 3'UTR that are present in the endogenous Arc gene. NMD depends on the presence of proteins of the exon junction complex (EJC) downstream of a stop codon, so EGFP-Arc mRNA should not undergo NMD. Assessment of Arc mRNA rundown in the presence of the transcription inhibitor actinomycin-D confirmed delayed degradation of EGFP-Arc mRNA. EGFP-Arc mRNA and protein are expressed at much higher levels in transgenic mice under basal and activated conditions but EGFP-Arc mRNA does not enter dendrites efficiently. In a physiological assay in which cycloheximide (CHX) was infused after induction of Arc by seizures, there were increases in endogenous Arc mRNA levels consistent with translation-dependent Arc mRNA decay but this was not seen with EGFP-Arc mRNA. Taken together, our results indicate: (1) Arc mRNA degradation occurs via a mechanism with characteristics of NMD; (2) rapid dendritic delivery of newly synthesized Arc mRNA after induction may depend in part on prior splicing of the 3'UTR.

Arc is a unique immediate early gene (IEG) whose expression is induced as synapses are modified during learning. Newly-synthesized Arc mRNA is rapidly transported throughout dendrites and localizes near recently activated synapses. Arc mRNA levels are regulated by rapid degradation, which is accelerated by synaptic activity in a translation-dependent process. One possible mechanism is nonsense-mediated mRNA decay (NMD), which depends on the presence of a splice junction in the 3'UTR. Here, we test this hypothesis using transgenic mice that express EGFP-Arc. Because the transgene was constructed from Arc cDNA, it lacks intron structures in the 3'UTR that are present in the endogenous Arc gene. NMD depends on the presence of proteins of the exon junction complex (EJC) downstream of a stop codon, so EGFP-Arc mRNA should not undergo NMD. Assessment of Arc mRNA rundown in the presence of the transcription inhibitor actinomycin-D confirmed delayed degradation of EGFP-Arc mRNA. EGFP-Arc mRNA and protein are expressed at much higher levels in transgenic mice under basal and activated conditions but EGFP-Arc mRNA does not enter dendrites efficiently. In a physiological assay in which cycloheximide (CHX) was infused after induction of Arc by seizures, there were increases in endogenous Arc mRNA levels consistent with translation-dependent Arc mRNA decay but this was not seen with EGFP-Arc mRNA. Taken together, our results indicate: (1) Arc mRNA degradation occurs via a mechanism with characteristics of NMD; (2) rapid dendritic delivery of newly synthesized Arc mRNA after induction may depend in part on prior splicing of the 3'UTR.

Abstract Background CpG islands are observed in mammals and other vertebrates, generally escape DNA methylation, and tend to occur in the promoters of widely expressed genes. Another class of promoter has lower G+C and CpG contents, and is thought to be involved in the spatiotemporal regulation of gene expression. Non-vertebrate deuterostomes are reported to have a single class of promoter with high-frequency CpG dinucleotides, suggesting that this is the original type of promoter. However, the limited annotation of these genes has impeded the large-scale analysis of their promoters. Results To determine the origins of the two classes of vertebrate promoters, we chose Ciona intestinalis, an invertebrate that is evolutionarily close to the vertebrates, and identified its transcription start sites genome-wide using a next-generation sequencer. We indeed observed a high CpG content around the transcription start sites, but their levels in the promoters and background sequences differed much less than in mammals. The CpG-rich stretches were also fairly restricted, so they appeared more similar to mammalian CpG-poor promoters. Conclusions From these data, we infer that CpG islands are not sufficiently ancient to be found in invertebrates. They probably appeared early in vertebrate evolution via some active mechanism and have since been maintained as part of vertebrate promoters.

Author(s): Richard Redon [1]; Shumpei Ishikawa [2, 3]; Karen R. Fitch [4]; Lars Feuk [5, 6]; George H. Perry [7]; T. Daniel Andrews [1]; Heike Fiegler [1]; Michael H. Shapero [...]

Abstract Background DNA methylation by the Dnmt family occurs in vertebrates and invertebrates, including ascidians, and is thought to play important roles in gene regulation and genome stability, especially in vertebrates. However, the global methylation patterns of vertebrates and invertebrates are distinctive. Whereas almost all CpG sites are methylated in vertebrates, with the exception of those in CpG islands, the ascidian genome contains approximately equal amounts of methylated and unmethylated regions. Curiously, methylation status can be reliably estimated from the local frequency of CpG dinucleotides in the ascidian genome. Methylated and unmethylated regions tend to have few and many CpG sites, respectively, consistent with our knowledge of the methylation status of CpG islands and other regions in mammals. However, DNA methylation patterns and levels in vertebrates and invertebrates have not been analyzed in the same way. Results Using a new computational methodology based on the decomposition of the bimodal distributions of methylated and unmethylated regions, we estimated the extent of the global methylation patterns in a wide range of animals. We then examined the epigenetic changes in silico along the phylogenetic tree. We observed a gradual transition from fractional to global patterns of methylation in deuterostomes, rather than a clear demarcation between vertebrates and invertebrates. When we applied this methodology to six piscine genomes, some of which showed features similar to those of invertebrates. Conclusions The mammalian global DNA methylation pattern was probably not acquired at an early stage of vertebrate evolution, but gradually expanded from that of a more ancient organism.

Summary: Peripheral T‐cell lymphoma (PTCL) is a group of heterogeneous non‐Hodgkin lymphomas showing a mature T‐cell or natural killer cell phenotype, but its molecular abnormalities in paediatric patients remain unclear. By employing next‐generation sequencing and multiplex ligation‐dependent probe amplification of tumour samples from 26 patients, we identified somatic alterations in paediatric PTCL including Epstein–Barr virus (EBV)‐negative (EBV–) and EBV‐positive (EBV+) patients. As recurrent mutational targets for PTCL, we identified several previously unreported genes, including TNS1, ZFHX3, LRP2,NCOA2 and HOXA1, as well as genes previously reported in adult patients, e.g. TET2, CDKN2A, STAT3 and TP53. However, for other reported mutations, VAV1‐related abnormalities were absent and mutations of NRAS, GATA3 and JAK3 showed a low frequency in our cohort. Concerning the association of EBV infection, two novel fusion genes: STAG2‐AFF2 and ITPR2‐FSTL4, and deletion and alteration of CDKN2A/2B, LMO1 and HOXA1 were identified in EBV– PTCL, but not in EBV+ PTCL. Conversely, alterations of PCDHGA4, ADAR, CUL9 and TP53 were identified only in EBV+ PTCL. Our observations suggest a clear difference in the molecular mechanism of onset between paediatric and adult PTCL and a difference in the characteristics of genetic alterations between EBV– and EBV+ paediatric PTCL. [ABSTRACT FROM AUTHOR]