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The INIA Texas Gene Expression Database: An online tool for alcohol genomics
Alcoholism is a serious condition that affects millions of people and costs billions of
dollars each year in treatment, damages, and lost income. In addition, it carries a
tremendous emotional burden. Alcoholism is caused by a combination of genetic
and environmental factors, which have yet to be fully identified. Fortunately,
alcoholism research, as well as research into other diseases with a genetic
component, has greatly benefited from recent rapid developments in
high-throughput genomic technologies and the development of relevant model
organisms. This has been highly productive for progress in the field, but effective
methods for identifying relevant data and for performing cross-dataset analyses
have not been developed at the same pace.
To help fulfill this need, I have developed the INIA (Integrative Neuroscience
Initiative on Alcoholism) Texas Gene Expression Database (IT-GED), which is freely
available at http://inia.icmb.utexas.edu. IT-GED is a web-based database which
contains a compilation of the significantly expressed genes from each of several
microarray datasets investigating the role of gene expression in the brain's
regulation of alcohol consumption. The studies were performed both in model
organisms (mouse and rat) and post-mortem humans. The data is presented via a
user-friendly interface which provides advanced searching abilities for identifying
genes of interest and tools for analysis of the data. These tools provide the ability to
compare user data to every dataset in IT-GED in order to assess the significance of
a group of genes across multiple datasets and the ability to generate visual
networks of those genes in order to identify the ones that are likely the most
functionally significant in the response to high alcohol consumption.
IT-GED thus provides a means by which alcohol researchers can combine multiple
sources of data to generate novel hypotheses concerning the genetic causes of
alcoholism. The goal of IT-GED is to provide support for comparing and integrating
results across gene expression studies of alcohol consumption and for generating
novel hypotheses based on individual genes and gene-gene interactions by
simplifying data access, providing various tools for analysis, and presenting users
with an easy-to-use interface.BiochemistryWaggoner Center for Alcohol and Addiction Researc
LIN28 Selectively Modulates a Subclass of Let-7 MicroRNAs
LIN28 is a bipartite RNA-binding protein that posttranscriptionallyinhibits the biogenesis of let-7 microRNAs to regulate development and influence disease states. However, the mechanisms of let-7 suppression remain poorly understood because LIN28 recognition depends on coordinated targeting by both the zinc knuckle domain (ZKD), which binds a GGAG-like element in the precursor, and the cold shock domain (CSD), whose binding sites have not been systematically characterized. By leveraging single-nucleotide-resolution mapping of LIN28 binding sites in vivo, we determined that the CSD recognizes a (U) GAU motif. This motif partitions the let-7 microRNAs into two subclasses, precursors with both CSD and ZKD binding sites (CSD+) and precursors with ZKD but no CSD binding sites (CSD-). LIN28 in vivo recognition-and subsequent 3' uridylation and degradation-of CSD+ precursors is more efficient, leading to their stronger suppression in LIN28-activated cells and cancers. Thus, CSD binding sites amplify the regulatory effects of LIN28
HITS-CLIP and Integrative Modeling Define the Rbfox Splicing-Regulatory Network Linked to Brain Development and Autism
The RNA binding proteins Rbfox1/2/3 regulate alternative splicing in the nervous system, and disruption of Rbfox1 has been implicated in autism. However, comprehensive identification of functional Rbfox targets has been challenging. Here, we perform HITS-CLIP for all three Rbfox family members in order to globally map, at a single-nucleotide resolution, their in vivo RNA interaction sites in the mouse brain. We find that the two guanines in the Rbfox binding motif UGCAUG are critical for protein-RNA interactions and crosslinking. Using integrative modeling, these interaction sites, combined with additional datasets, define 1,059 direct Rbfox target alternative splicing events. Over half of the quantifiable targets show dynamic changes during brain development. Of particular interest are 111 events from 48 candidate autism-susceptibility genes, including syndromic autism genes Shank3, Cacna1c, and Tsc2. Alteration of Rbfox targets in some autistic brains is correlated with downregulation of all three Rbfox proteins, supporting the potential clinical relevance of the splicing-regulatory network. © 2014 The Authors
Antisense oligonucleotide modulation of non-productive alternative splicing upregulates gene expression
Restoration of normal gene expression is one way to treat monogenic disorders. Here the authors target naturally occurring non-productive alternative splicing using antisense oligonucleotides to promote the production of functional proteins
MBNL Sequestration by Toxic RNAs and RNA Misprocessing in the Myotonic Dystrophy Brain
For some neurological disorders, disease is primarily RNA mediated due to expression of non-coding microsatellite expansion RNAs (RNAexp). Toxicity is thought to result from enhanced binding of proteins to these expansions and depletion from their normal cellular targets. However, experimental evidence for this sequestration model is lacking. Here, we use HITS-CLIP and pre-mRNA processing analysis of human control versus myotonic dystrophy (DM) brains to provide compelling evidence for this RNA toxicity model. MBNL2 binds directly to DM repeat expansions in the brain, resulting in depletion from its normal RNA targets with downstream effects on alternative splicing and polyadenylation. Similar RNA processing defects were detected in Mbnl compound-knockout mice, highlighted by dysregulation of Mapt splicing and fetal tau isoform expression in adults. These results demonstrate that MBNL proteins are directly sequestered by RNAexp in the DM brain and introduce a powerful experimental tool to evaluate RNA-mediated toxicity in other expansion diseases