Subcellular relocalization and nuclear redistribution of the RNA methyltransferases TRMT1 and TRMT1L upon neuronal activation.

RNA modifications are dynamic chemical entities that expand the RNA lexicon and regulate RNA fate. The most abundant modification present in mRNAs, N6-methyladenosine (m6A), has been implicated in neurogenesis and memory formation. However, whether additional RNA modifications may be playing a role in neuronal functions and in response to environmental queues is largely unknown. Here we characterize the biochemical function and cellular dynamics of two human RNA methyltransferases previously associated with neurological dysfunction, TRMT1 and its homolog, TRMT1-like (TRMT1L). Using a combination of next-generation sequencing, LC-MS/MS, patient-derived cell lines and knockout mouse models, we confirm the previously reported dimethylguanosine (m2,2G) activity of TRMT1 in tRNAs, as well as reveal that TRMT1L, whose activity was unknown, is responsible for methylating a subset of cytosolic tRNAAla(AGC) isodecoders at position 26. Using a cellular in vitro model that mimics neuronal activation and long term potentiation, we find that both TRMT1 and TRMT1L change their subcellular localization upon neuronal activation. Specifically, we observe a major subcellular relocalization from mitochondria and other cytoplasmic domains (TRMT1) and nucleoli (TRMT1L) to different small punctate compartments in the nucleus, which are as yet uncharacterized. This phenomenon does not occur upon heat shock, suggesting that the relocalization of TRMT1 and TRMT1L is not a general reaction to stress, but rather a specific response to neuronal activation. Our results suggest that subcellular relocalization of RNA modification enzymes may play a role in neuronal plasticity and transmission of information, presumably by addressing new targets.

The RNA modification landscape in human disease.

RNA modifications have been historically considered as fine-tuning chemo-structural features of infrastructural RNAs, such as rRNAs, tRNAs, and snoRNAs. This view has changed dramatically in recent years, to a large extent as a result of systematic efforts to map and quantify various RNA modifications in a transcriptome-wide manner, revealing that RNA modifications are reversible, dynamically regulated, far more widespread than originally thought, and involved in major biological processes, including cell differentiation, sex determination, and stress responses. Here we summarize the state of knowledge and provide a catalog of RNA modifications and their links to neurological disorders, cancers, and other diseases. With the advent of direct RNA-sequencing technologies, we expect that this catalog will help prioritize those RNA modifications for transcriptome-wide maps.

Adar3 Is Involved in Learning and Memory in Mice.

The amount of regulatory RNA encoded in the genome and the extent of RNA editing by the post-transcriptional deamination of adenosine to inosine (A-I) have increased with developmental complexity and may be an important factor in the cognitive evolution of animals. The newest member of the A-I editing family of ADAR proteins, the vertebrate-specific ADAR3, is highly expressed in the brain, but its functional significance is unknown. In vitro studies have suggested that ADAR3 acts as a negative regulator of A-I RNA editing but the scope and underlying mechanisms are also unknown. Meta-analysis of published data indicates that mouse Adar3 expression is highest in the hippocampus, thalamus, amygdala, and olfactory region. Consistent with this, we show that mice lacking exon 3 of Adar3 (which encodes two double stranded RNA binding domains) have increased levels of anxiety and deficits in hippocampus-dependent short- and long-term memory formation. RNA sequencing revealed a dysregulation of genes involved in synaptic function in the hippocampi of Adar3-deficient mice. We also show that ADAR3 transiently translocates from the cytoplasm to the nucleus upon KCl-mediated activation in SH-SY5Y cells. These results indicate that ADAR3 contributes to cognitive processes in mammals.

The long non-coding RNA NEAT1 is responsive to neuronal activity and is associated with hyperexcitability states.

Despite their abundance, the molecular functions of long non-coding RNAs in mammalian nervous systems remain poorly understood. Here we show that the long non-coding RNA, NEAT1, directly modulates neuronal excitability and is associated with pathological seizure states. Specifically, NEAT1 is dynamically regulated by neuronal activity in vitro and in vivo, binds epilepsy-associated potassium channel-interacting proteins including KCNAB2 and KCNIP1, and induces a neuronal hyper-potentiation phenotype in iPSC-derived human cortical neurons following antisense oligonucleotide knockdown. Next generation sequencing reveals a strong association of NEAT1 with increased ion channel gene expression upon activation of iPSC-derived neurons following NEAT1 knockdown. Furthermore, we show that while NEAT1 is acutely down-regulated in response to neuronal activity, repeated stimulation results in NEAT1 becoming chronically unresponsive in independent in vivo rat model systems relevant to temporal lobe epilepsy. We extended previous studies showing increased NEAT1 expression in resected cortical tissue from high spiking regions of patients suffering from intractable seizures. Our results indicate a role for NEAT1 in modulating human neuronal activity and suggest a novel mechanistic link between an activity-dependent long non-coding RNA and epilepsy.

Seq and You Will Find.

The human genome sequence is freely available, nearly complete and is providing a foundation of research opportunities that are overturning our current understanding of human biology. The advent of next generation sequencing has revolutionized the way we can interrogate the genome and its transcriptional products and how we analyze, diagnose, monitor and even treat human disease. Personal genetic profiles are increasing dramatically in medical value as researchers accumulate more and more knowledge about the interaction between genetic and environmental factors that contribute to the onset of common disorders. As the cost of sequencing plummets, whole genome sequencing of individuals is becoming a reality and the field of personalized genomic medicine is rapidly developing. Now there is great need for accurate annotation of all functionally important sequences in the human genome and the variations within them that contribute to health and disease. The vast majority of our genome gives rise to RNA transcripts. This extraordinarily versatile molecule not only encodes protein information but also has great structural dynamics and plasticity, capacity for DNA/RNA/protein interactions and catalytic activity. It is a key regulator of biological networks with clear links to human disease and a more comprehensive understanding of its function is needed to maximise its use in medical practice. This review focuses on the complexity of our genome and the impact of sequencing technologies in understanding its many products and functions in health and disease.