Widespread splicing of repetitive element loci into coding regions of gene transcripts.

We performed a thorough characterization of expressed repetitive element loci (RE) in the human orbitofrontal cortex (OFC) using directional RNA sequencing data. Considering only sequencing reads that map uniquely onto the human genome, we discovered that the overwhelming majority of intronic and exonic RE are expressed in the same orientation as the gene in which they reside. Our mapping approach enabled the identification of novel differentially expressed RE transcripts between the OFC and peripheral blood lymphocytes. Further analysis revealed that RE are extensively spliced into coding regions of gene transcripts yielding thousands of novel mRNA variants with altered coding potential. Lower frequency splicing of RE into untranslated regions of gene transcripts was also observed. The same pattern of RE splicing in the brain was also detected for Drosophila, zebrafish, mouse, rat, dog and rabbit. RE splicing occurs largely at canonical GT-AG splice junctions with LINE and SINE elements forming the most RE splice junctions in the human OFC. This type of splicing usually gives rise to a minor splice variant of the endogenous gene and in silico analysis suggests that RE splicing has the potential to introduce novel open reading frames. Reanalysis of previously published sequencing data performed in the mouse cerebellum revealed that thousands of RE splice variants are associated with translating ribosomes. Our results demonstrate that RE expression is more complex than previously envisioned and raise the possibility that RE splicing might generate functional protein isoforms.

Repetitive elements and epigenetic marks in behavior and psychiatric disease.

Repetitive elements, which are relics of previous transposition events, constitute a large proportion of the human genome. The ability of transposons to gives rise to new DNA combinations has clearly provided an evolutionary advantage to their hosts. Transposons have shaped our genomes by giving rise to novel coding sequences, alternative gene promoters, conserved noncoding elements, and gene networks. Despite its benefits, the process of transposition can also create deleterious DNA combinations, and a growing number of human diseases are being linked to abnormal repetitive element activity. To limit transposition, cells tightly regulate and immobilize repetitive elements using DNA methylation and other epigenetic marks. Recent findings in neuropsychiatric disorders implicate both repetitive elements and epigenetic marks as potential etiological factors. It is possible that these observations are linked and that the reported alterations in epigenetic marks may create a permissive state enabling transposons to mobilize. In this work, we provide a detailed description of repetitive element biology and epigenetics to familiarize the readers with the subject matter and to illustrate how their disruption can result in pathology. We also review the evidence for the involvement of these two factors in neuropsychiatric disorders and discuss the need for replication studies to confirm these initial findings. We are cautiously optimistic that further characterization of epigenetic mark and repetitive element activity in the brain will reveal the underlying causes of schizophrenia, bipolar disorder, and major depression.

The Saccharomyces cerevisiae Nrd1-Nab3 transcription termination pathway acts in opposition to Ras signaling and mediates response to nutrient depletion.

The Saccharomyces cerevisiae Nrd1-Nab3 pathway directs the termination and processing of short RNA polymerase II transcripts. Despite the potential for Nrd1-Nab3 to affect the transcription of both coding and noncoding RNAs, little is known about how the Nrd1-Nab3 pathway interacts with other pathways in the cell. Here we present the results of a high-throughput synthetic lethality screen for genes that interact with NRD1 and show roles for Nrd1 in the regulation of mitochondrial abundance and cell size. We also provide genetic evidence of interactions between the Nrd1-Nab3 and Ras/protein kinase A (PKA) pathways. Whereas the Ras pathway promotes the transcription of genes involved in growth and glycolysis, the Nrd1-Nab3 pathway appears to have a novel role in the rapid suppression of some genes when cells are shifted to poor growth conditions. We report the identification of new mRNA targets of the Nrd1-Nab3 pathway that are rapidly repressed in response to glucose depletion. Glucose depletion also leads to the dephosphorylation of Nrd1 and the formation of novel nuclear speckles that contain Nrd1 and Nab3. Taken together, these results indicate a role for Nrd1-Nab3 in regulating the cellular response to nutrient availability.

Transcriptome-wide binding sites for components of the Saccharomyces cerevisiae non-poly(A) termination pathway: Nrd1, Nab3, and Sen1.

RNA polymerase II synthesizes a diverse set of transcripts including both protein-coding and non-coding RNAs. One major difference between these two classes of transcripts is the mechanism of termination. Messenger RNA transcripts terminate downstream of the coding region in a process that is coupled to cleavage and polyadenylation reactions. Non-coding transcripts like Saccharomyces cerevisiae snoRNAs terminate in a process that requires the RNA-binding proteins Nrd1, Nab3, and Sen1. We report here the transcriptome-wide distribution of these termination factors. These data sets derived from in vivo protein-RNA cross-linking provide high-resolution definition of non-poly(A) terminators, identify novel genes regulated by attenuation of nascent transcripts close to the promoter, and demonstrate the widespread occurrence of Nrd1-bound 3' antisense transcripts on genes that are poorly expressed. In addition, we show that Sen1 does not cross-link efficiently to many expected non-coding RNAs but does cross-link to the 3' end of most pre-mRNA transcripts, suggesting an extensive role in mRNA 3' end formation and/or termination.

Yeast Nrd1, Nab3, and Sen1 transcriptome-wide binding maps suggest multiple roles in post-transcriptional RNA processing.

RNA polymerase II transcribes both coding and noncoding genes, and termination of these different classes of transcripts is facilitated by different sets of termination factors. Pre-mRNAs are terminated through a process that is coupled to the cleavage/polyadenylation machinery, and noncoding RNAs in the yeast Saccharomyces cerevisiae are terminated through a pathway directed by the RNA-binding proteins Nrd1, Nab3, and the RNA helicase Sen1. We have used an in vivo cross-linking approach to map the binding sites of components of the yeast non-poly(A) termination pathway. We show here that Nrd1, Nab3, and Sen1 bind to a number of noncoding RNAs in an unexpected manner. Sen1 shows a preference for H/ACA over box C/D snoRNAs. Nrd1, which binds to snoRNA terminators, also binds to the upstream region of some snoRNA transcripts and to snoRNAs embedded in introns. We present results showing that several RNAs, including the telomerase RNA TLC1, require Nrd1 for proper processing. Binding of Nrd1 to transcripts from tRNA genes is another unexpected observation. We also observe RNA polymerase II binding to transcripts from RNA polymerase III genes, indicating a possible role for the Nrd1 pathway in surveillance of transcripts synthesized by the wrong polymerase. The binding targets of Nrd1 pathway components change in the absence of glucose, with Nrd1 and Nab3 showing a preference for binding to sites in the mature snoRNA and tRNAs. This suggests a novel role for Nrd1 and Nab3 in destruction of ncRNAs in response to nutrient limitation.