Joel R. Neilson

Bio: Joel R. Neilson is an academic researcher from Baylor College of Medicine. The author has contributed to research in topics: Calcineurin & NFAT. The author has an hindex of 35, co-authored 63 publications receiving 10464 citations. Previous affiliations of Joel R. Neilson include Baylor University & Massachusetts Institute of Technology.

MicroRNA sponges: competitive inhibitors of small RNAs in mammalian cells

Abstract: MicroRNAs are predicted to regulate thousands of mammalian genes, but relatively few targets have been experimentally validated and few microRNA loss-of-function phenotypes have been assigned. As an alternative to chemically modified antisense oligonucleotides, we developed microRNA inhibitors that can be expressed in cells, as RNAs produced from transgenes. Termed 'microRNA sponges', these competitive inhibitors are transcripts expressed from strong promoters, containing multiple, tandem binding sites to a microRNA of interest. When vectors encoding these sponges are transiently transfected into cultured cells, sponges derepress microRNA targets at least as strongly as chemically modified antisense oligonucleotides. They specifically inhibit microRNAs with a complementary heptameric seed, such that a single sponge can be used to block an entire microRNA seed family. RNA polymerase II promoter (Pol II)-driven sponges contain a fluorescence reporter gene for identification and sorting of sponge-treated cells. We envision the use of stably expressed sponges in animal models of disease and development.

Proliferating Cells Express mRNAs with Shortened 3' Untranslated Regions and Fewer MicroRNA Target Sites

Abstract: Messenger RNA (mRNA) stability, localization, and translation are largely determined by sequences in the 3' untranslated region (3'UTR). We found a conserved increase in expression of mRNAs terminating at upstream polyadenylation sites after activation of primary murine CD4+ T lymphocytes. This program, resulting in shorter 3'UTRs, is a characteristic of gene expression during immune cell activation and correlates with proliferation across diverse cell types and tissues. Forced expression of full-length 3'UTRs conferred reduced protein expression. In some cases the reduction in protein expression could be reversed by deletion of predicted microRNA target sites in the variably included region. Our data indicate that gene expression is coordinately regulated, such that states of increased proliferation are associated with widespread reductions in the 3'UTR-based regulatory capacity of mRNAs.

NFAT dysregulation by increased dosage of DSCR1 and DYRK1A on chromosome 21

Abstract: Trisomy 21 results in Down's syndrome, but little is known about how a 1.5-fold increase in gene dosage produces the pleiotropic phenotypes of Down's syndrome. Here we report that two genes, DSCR1 and DYRK1A , lie within the critical region of human chromosome 21 and act synergistically to prevent nuclear occupancy of NFATc transcription factors, which are regulators of vertebrate development. We use mathematical modelling to predict that autoregulation within the pathway accentuates the effects of trisomy of DSCR1 and DYRK1A, leading to failure to activate NFATc target genes under specific conditions. Our observations of calcineurin-and Nfatc-deficient mice, Dscr1- and Dyrk1a-overexpressing mice, mouse models of Down's syndrome and human trisomy 21 are consistent with these predictions. We suggest that the 1.5-fold increase in dosage of DSCR1 and DYRK1A cooperatively destabilizes a regulatory circuit, leading to reduced NFATc activity and many of the features of Down's syndrome. More generally, these observations suggest that the destabilization of regulatory circuits can underlie human disease.

L-type calcium channels and GSK-3 regulate the activity of NF-ATc4 in hippocampal neurons

Abstract: The molecular basis of learning and memory has been the object of several recent advances, which have focused attention on calcium-regulated pathways controlling transcription. One of the molecules implicated by pharmacological, biochemical and genetic approaches is the calcium/calmodulin-regulated phosphatase, calcineurin. In lymphocytes, calcineurin responds to specific calcium signals and regulates expression of several immediate early genes by controlling the nuclear import of the NF-ATc family of transcription factors. Here we show that NF-ATc4/NF-AT3 in hippocampal neurons can rapidly translocate from cytoplasm to nucleus and activate NF-AT-dependent transcription in response to electrical activity or potassium depolarization. The calcineurin-mediated translocation is critically dependent on calcium entry through L-type voltage-gated calcium channels. GSK-3 can phosphorylate NF-ATc4, promoting its export from the nucleus and antagonizing NF-ATc4-dependent transcription. Furthermore, we show that induction of the inositol 1,4,5-trisphosphate receptor type 1 is controlled by the calcium/calcineurin/NF-ATc pathway. This provides a new perspective on the function of calcineurin in the central nervous system and indicates that NF-AT-mediated gene expression may be involved in the induction of hippocampal synaptic plasticity and memory formation.

Circular RNAs are a large class of animal RNAs with regulatory potency

Abstract: Circular RNAs (circRNAs) in animals are an enigmatic class of RNA with unknown function. To explore circRNAs systematically, we sequenced and computationally analysed human, mouse and nematode RNA. We detected thousands of well-expressed, stable circRNAs, often showing tissue/developmental-stage-specific expression. Sequence analysis indicated important regulatory functions for circRNAs. We found that a human circRNA, antisense to the cerebellar degeneration-related protein 1 transcript (CDR1as), is densely bound by microRNA (miRNA) effector complexes and harbours 63 conserved binding sites for the ancient miRNA miR-7. Further analyses indicated that CDR1as functions to bind miR-7 in neuronal tissues. Human CDR1as expression in zebrafish impaired midbrain development, similar to knocking down miR-7, suggesting that CDR1as is a miRNA antagonist with a miRNA-binding capacity ten times higher than any other known transcript. Together, our data provide evidence that circRNAs form a large class of post-transcriptional regulators. Numerous circRNAs form by head-to-tail splicing of exons, suggesting previously unrecognized regulatory potential of coding sequences.

Natural RNA circles function as efficient microRNA sponges

Abstract: MicroRNAs (miRNAs) are important post-transcriptional regulators of gene expression that act by direct base pairing to target sites within untranslated regions of messenger RNAs. Recently, miRNA activity has been shown to be affected by the presence of miRNA sponge transcripts, the so-called competing endogenous RNA in humans and target mimicry in plants. We previously identified a highly expressed circular RNA (circRNA) in human and mouse brain. Here we show that this circRNA acts as a miR-7 sponge; we term this circular transcript ciRS-7 (circular RNA sponge for miR-7). ciRS-7 contains more than 70 selectively conserved miRNA target sites, and it is highly and widely associated with Argonaute (AGO) proteins in a miR-7-dependent manner. Although the circRNA is completely resistant to miRNA-mediated target destabilization, it strongly suppresses miR-7 activity, resulting in increased levels of miR-7 targets. In the mouse brain, we observe overlapping co-expression of ciRS-7 and miR-7, particularly in neocortical and hippocampal neurons, suggesting a high degree of endogenous interaction. We further show that the testis-specific circRNA, sex-determining region Y (Sry), serves as a miR-138 sponge, suggesting that miRNA sponge effects achieved by circRNA formation are a general phenomenon. This study serves as the first, to our knowledge, functional analysis of a naturally expressed circRNA.

Predicting effective microRNA target sites in mammalian mRNAs

Abstract: Proteins are built by using the information contained in molecules of messenger RNA (mRNA). Cells have several ways of controlling the amounts of different proteins they make. For example, a so-called ‘microRNA’ molecule can bind to an mRNA molecule to cause it to be more rapidly degraded and less efficiently used, thereby reducing the amount of protein built from that mRNA. Indeed, microRNAs are thought to help control the amount of protein made from most human genes, and biologists are working to predict the amount of control imparted by each microRNA on each of its mRNA targets. All RNA molecules are made up of a sequence of bases, each commonly known by a single letter—‘A’, ‘U’, ‘C’ or ‘G’. These bases can each pair up with one specific other base—‘A’ pairs with ‘U’, and ‘C’ pairs with ‘G’. To direct the repression of an mRNA molecule, a region of the microRNA known as a ‘seed’ binds to a complementary sequence in the target mRNA. ‘Canonical sites’ are regions in the mRNA that contain the exact sequence of partner bases for the bases in the microRNA seed. Some canonical sites are more effective at mRNA control than others. ‘Non-canonical sites’ also exist in which the pairing between the microRNA seed and mRNA does not completely match. Previous work has suggested that many non-canonical sites can also control mRNA degradation and usage. Agarwal et al. first used large experimental datasets from many sources to investigate microRNA activity in more detail. As expected, when mRNAs had canonical sites that matched the microRNA, mRNA levels and usage tended to drop. However, no effect was observed when the mRNAs only had recently identified non-canonical sites. This suggests that microRNAs primarily bind to canonical sites to control protein production. Based on these results, Agarwal et al. further developed a statistical model that predicts the effects of microRNAs binding to canonical sites. The updated model considers 14 different features of the microRNA, microRNA site, or mRNA—including the mRNA sequence around the site—to predict which sites within mRNAs are most effectively targeted by microRNAs. Tests showed that Agarwal et al.'s model was as good as experimental approaches at identifying the effective target sites, and was better than existing computational models. The model has been used to power the latest version of a freely available resource called TargetScan, and so could prove a valuable resource for researchers investigating the many important roles of microRNAs in controlling protein production.