Ongoing projects

The Berglund lab continues to focus on microsatellite diseases with the goal of translating basic science into clinical research using a combination of biochemical, cellular and genomic approaches.  As potential new therapies for neuromuscular and neurodegenerative diseases progress through preclinical and clinical evaluation, drug developers need new tools to design and conduct clinical trials. Research in the Berglund lab focuses on the identification of potential biomarkers and potential therapeutic molecules for microsatellite diseases such as myotonic dystrophy types 1 and 2 (DM1 and DM2), spinocerebellar ataxias (SCAs), and amyotrophic lateral sclerosis (ALS).

 

RNA Splicing and Disease

The Berglund lab uses a broad range of approaches to study the molecular mechanisms of neurological diseases that are caused by microsatellite repeat expansions.  For many of these diseases (DM, ALS and SCAs), RNA processing (pre-mRNA splicing) pathways are negatively impacted with specific changes in pre-mRNA splicing proposed to lead to symptoms observed in affected individuals. Many of the projects in the lab combine biochemical, molecular, and genomic approaches with cellular and other model systems to understand the mechanisms through which these diseases alter pre-mRNA splicing. In one project we have developed a framework to estimate MBNL concentration using splicing responses alone and validated it in a cell-based model and then applied it to myotonic dystrophy patient tissue. This allowed us to evaluate the ability of individual and combinations of splicing events from genomic data to predict functional MBNL concentration in human biopsies and the its potential use as a biomarker to distinguish mild, moderate, and severe cases of DM.

 

Dose-Dependent Regulation of Alternative Splicing by MBNL Proteins Reveals Biomarkers for Myotonic Dystrophy. Wagner SD, Struck AJ, Gupta R, Farnsworth DR, Mahady AE, Eichinger K, Thornton CA, Wang ET, Berglund JA. PLoS Genet. 2016 Sep 28;12(9):e1006316. doi: 10.1371/journal.pgen.1006316. eCollection 2016 Sep. PMID: 27681373

 

Small Molecule Targeting of toxic RNA

We are developing approaches to screen libraries of small molecules and mining the scientific literature to identify compounds that can be used to inhibit the production of toxic RNAs. Lead compounds that show promise inhibiting the production of the toxic RNAs will be studied further to understand the mechanisms through which they function.  This could provide fundamental information for the development of molecules with improved activity.  The goal of our research is to use the results from these fundamental studies to identify innovative strategies to reduce or correct the improper pre-mRNA splicing that occurs in the disease state. For example, we have recently shown that small molecules can be used to rescue the mis-splicing in cell and mouse models of myotonic dystrophy.  Our investigation into the mode of action of furamidine, a promising small molecule for rescue of mis-splicing in DM1 cells, showed that furamidine affects multiple pathways of DM1 pathogenesis and that it may work through multiple mechanisms to rescue DM1-associated mis-splicing events.

 

Combination Treatment of Erythromycin and Furamidine Provides Additive and Synergistic Rescue of Mis-splicing in Myotonic Dystrophy Type 1 Models Jenquin, J.R., Yang, H., Huigens, R.W., Nakamori, M., Berglund, A. ACS Pharmacol. Transl. Sci. · July 17, 2019

 

Furamidine Rescues Myotonic Dystrophy Type I Associated Mis-Splicing through Multiple Mechanisms. Jenquin JR, Coonrod LA, Silverglate QA, Pellitier NA, Hale MA, Xia G, Nakamori M, Berglund JA. ACS Chem Biol. 2018 Sep 21;13(9):2708-2718. doi: 10.1021/acschembio.8b00646. Epub 2018 Aug 27. PMID: 30118588

 

Engineering Proteins

We are developing novel synthetic muscleblind-like 1 (MBNL1) proteins, with altered and new activities, to provide insight into how this protein recognizes RNA and regulates splicing. We have recently published some of this work in Nucleic Acid Research describing an engineered MBNL1 RNA binding protein with improved specificity.  We showed that MBNL1’s zinc finger (ZF) domains have different activities with the first domain ZF1-2 driving splicing regulation while ZF3-4 acts as a general RNA binding domain. These studies suggest that synthetic MBNL proteins with altered splicing activity have the potential to be used as both tools for investigating splicing regulation and as potential protein therapeutics for DM and other repeat disease.

 

An engineered RNA binding protein with improved splicing regulation. Hale MA, Richardson JI, Day RC, McConnell OL, Arboleda J, Wang ET, Berglund JA. Nucleic Acids Res. 2018 Apr 6;46(6):3152-3168. doi: 10.1093/nar/gkx1304. PMID:  29309648

 

Genomics

RNA-seq is a powerful tool that is used to study the transcriptome. We use RNA-seq to compare the transcriptomes of DM models (cell and animal) as well as to determine how small molecules or how loss or gain of RNA binding proteins affect the transcriptome in these DM models. For example, in our recent study with furamidine we examined the global splicing and gene expression changes caused by furamidine in a DM1 mouse model. We were excited to observe that furamidine had significantly fewer off-target effects on the transcriptome compared to Actinomycin D, a small molecule that we had studied previously.

 

Furamidine Rescues Myotonic Dystrophy Type I Associated Mis-Splicing through Multiple Mechanisms. Jenquin JR, Coonrod LA, Silverglate QA, Pellitier NA, Hale MA, Xia G, Nakamori M, Berglund JA. ACS Chem Biol. 2018 Sep 21;13(9):2708-2718. doi: 10.1021/acschembio.8b00646. Epub 2018 Aug 27. PMID: 30118588

 

Models of Disease

For many years our group has used cell and animal models to identify small molecules to target the toxic CUG and CCUG repeats of DM in an effort to identify molecules that rescue the mis-splicing in DM. In addition to mouse models and disease specific human cell lines we have been collaborating with Dr. Karen Guillemin’s lab at the University of Oregon on developing and characterizing zebrafish models of myotoinic dystrophy.  The main focus of the zebrafish project is to the study the mechanisms underlying DM-related changes in gut motility and the microbiome since this has been a common issue among DM patients.

 

Modifications to toxic CUG RNAs induce structural stability, rescue mis-splicing in a myotonic dystrophy cell model and reduce toxicity in a myotonic dystrophy zebrafish model. deLorimier E, Coonrod LA, Copperman J, Taber A, Reister EE, Sharma K, Todd PK, Guenza MG, Berglund JA. Nucleic Acids Res. 2014 Nov 10;42(20):12768-78. doi: 10.1093/nar/gku941. Epub 2014 Oct 10. PMID: 25303993

 

RNA Structure

One of the mechanisms through which the CUG repeats and CCUG repeats (for myotonic dystrophy type 1 (DM1) and 2 (DM2) respectively) are toxic, is the sequestration of the muscleblind-like (MBNL) family of RNA binding proteins. The sequestration of MBNL proteins leads to many changes in splicing, which are implicated in causing the symptoms in DM.  It is therefore important to understand how MBNL proteins bind to their toxic and cellular RNA substrates to develop mechanisms to alleviate MBNL sequestration in DM1 and DM2.  MBNL proteins bind to a consensus YGCY RNA sequence and since CUG and CCUG expansion repeats are composed of YGCY motifs there are potentially hundreds to thousands of MBNL-binding sites in someone with a DM1 or DM2 expansion. Our lab has published work on replacing uridines with pseudouridine which induced structural stabilization, prevented MBNL1 binding, and rescued mis-splicing.  We continue to use crystallography and structure probing methods to better understand the structure of RNA in the context of DM and other repeat diseases.

 

Pseudouridine Modification Inhibits Muscleblind-like 1 (MBNL1) Binding to CCUG Repeats and Minimally Structured RNA through Reduced RNA Flexibility. deLorimier E, Hinman MN, Copperman J, Datta K, Guenza M, Berglund JA. J Biol Chem. 2017 Mar 10;292(10):4350-4357. doi: 10.1074/jbc.M116.770768. Epub 2017 Jan 27. PMID: 28130447

 

Utilizing the GAAA tetraloop/receptor to facilitate crystal packing and determination of the structure of a CUG RNA helix. Coonrod LA, Lohman JR, Berglund JA. Biochemistry. 2012 Oct 23;51(42):8330-7. doi: 10.1021/bi300829w. Epub 2012 Oct 12. PMID: 23025897


Publications

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Recent Publications

Lab Members

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