Research Highlights

A brief look at the latest discoveries from RBI faculty and trainees.

ENTPD3 marks mature stem cell derived beta cells formed by self-aggregation in vitro

August 11, 2021

Fiona M Docherty, Kent A Riemondy, Roberto Castro-Gutierrez, JaeAnn M Dwulet, Ali H Shilleh, Maria S Hansen, Shane P M Williams, Lucas H Armitage, Katherine E Santostefano, Mark A Wallet, Clayton E Mathews, Taylor M Triolo, Richard K P Benninger, Holger A Russ

Stem cell derived beta-like cells (sBC) carry the promise of providing an abundant source of insulin-producing cells for use in cell replacement therapy for patients with diabetes, potentially allowing widespread implementation of a practical cure. To achieve their clinical promise, sBC need to function comparably to mature adult beta cells, but as yet they display varying degrees of maturity. Complete Abstract

Srsf3 mediates alternative RNA splicing downstream of PDGFRα signaling in the facial mesenchyme

July 15, 2021

Brenna J C Dennison, Eric D Larson, Rui Fu, Julia Mo, Katherine A Fantauzzo

Signaling through the platelet-derived growth factor receptor alpha (PDGFRα) is crucial for mammalian craniofacial development, although the mechanisms by which the activity of downstream intracellular effectors is regulated to mediate gene expression changes have not been defined. We find that the RNA-binding protein Srsf3 is phosphorylated at Akt consensus sites downstream of PI3K-mediated PDGFRα signaling in mouse palatal mesenchyme cells, leading to its nuclear translocation. Complete Abstract

LABRAT reveals association of alternative polyadenylation with transcript localization, RNA binding protein expression, transcription speed, and cancer survival

June 26, 2021

Raeann Goering, Krysta L Engel, Austin E Gillen, Nova Fong, David L Bentley, J Matthew Taliaferro  

The sequence content of the 3' UTRs of many mRNA transcripts is regulated through alternative polyadenylation (APA). The study of this process using RNAseq data, though, has been historically challenging. Complete Abstract

Three-dimensional structure of a flavivirus dumbbell RNA reveals molecular details of an RNA regulator of replication

June 16, 2021

Benjamin M Akiyama, Monica E Graham, Zoe O Donoghue, J David Beckham, Jeffrey S Kieft  

Mosquito-borne flaviviruses (MBFVs) including dengue, West Nile, yellow fever, and Zika viruses have an RNA genome encoding one open reading frame flanked by 5' and 3' untranslated regions (UTRs). The 3' UTRs of MBFVs contain regions of high sequence conservation in structured RNA elements known as dumbbells (DBs). Complete Abstract

RNA-binding proteins regulate aldosterone homeostasis in human steroidogenic cells

June 1, 2021

Rui Fu, Kimberly Wellman, Amber Baldwin, Juilee Rege, Kathryn Walters, Antje Hirsekorn, Kent Riemondy, William E Rainey and Neelanjan Mukherjee

Angiotensin II (AngII) stimulates adrenocortical cells to produce aldosterone, a master regulator of blood pressure. Despite extensive characterization of the transcriptional and enzymatic control of adrenocortical steroidogenesis, there are still major gaps in the precise regulation of AII-induced gene expression kinetics. Complete Abstract

Cell-level metadata are indispensable for documenting single-cell sequencing datasets

May 24, 2021

Sidhant Puntambekar, Jay R Hesselberth, Kent A Riemondy, Rui Fu 

Single-cell RNA sequencing (scRNA-seq) has empowered discoveries of cell heterogeneity and state transitions at unprecedented resolution and throughput. New technological developments have broadened the scope of measurable molecules, extending beyond RNA to measure cell surface proteins. Complete Abstract

Establishing RNA-RNA Interactions Remodels lncRNA Structure and Promotes PRC2 Activity  

April 14, 2021

Maggie M. Balas, Erik W. Hartwick, Chloe Barrington, Justin T. Roberts, Stephen K. Wu, Ryan Bettcher, April M. Griffin, Jeffrey S. Kieft, Aaron M. Johnson

Human Polycomb Repressive Complex 2 (PRC2) catalysis of histone H3 lysine 27 methylation at certain loci depends on long noncoding RNAs (lncRNAs). Yet, in apparent contradiction, RNA is a potent catalytic inhibitor of PRC2. Here, we show that intermolecular RNA-RNA interactions between the lncRNA HOTAIR and its targets can relieve RNA inhibition of PRC2. Complete Abstract

Molecular Tracking Devices Quantify Antigen Distribution and Archiving in the Murine Lymph Node

April 12, 2021

Shannon M Walsh, Ryan M Sheridan, Erin D Lucas, Thu A Doan, Brian C Ware, Johnathon Schafer, Rui Fu, Matthew A Burchill, Jay R Hesselberth, Beth Ann Jiron Tamburini

The detection of foreign antigens in vivo has relied on fluorescent conjugation or indirect read-outs such as antigen presentation. In our studies, we found that these widely used techniques had several technical limitations that have precluded a complete picture of antigen trafficking or retention across lymph node cell types.  Complete Abstract

ELAV/Hu RNA Binding Proteins Determine Multiple Programs of Neural Alternative Splicing

April 7, 2021

Seungjae Lee, Lu Wei, Binglong Zhang, Raeann Goering, Sonali Majumdar, Jiayu Wen, J Matthew Taliaferro, Eric C Lai 

ELAV/Hu factors are conserved RNA binding proteins (RBPs) that play diverse roles in mRNA processing and regulation. The founding member, Drosophila Elav, was recognized as a vital neural factor 35 years ago. Nevertheless, little was known about its impact on the transcriptome, and potential functional overlap with its paralogs. Complete Abstract

Alternative RNA Structures Formed During Transcription Depend on Elongation Rate and Modify RNA Processing

February 24, 2021

Tassa Saldi, Kent Riemondy, Benjamin Erickson, and David L. Bentley

Most RNA processing occurs co-transcriptionally. We interrogated nascent pol II transcripts by chemical and enzymatic probing and determined how the “nascent RNA structureome” relates to splicing, A-I editing and transcription speed. RNA folding within introns and steep structural transitions at splice sites are associated with efficient co-transcriptional splicing. Complete Abstract

Qualitative Differences Between the IFNα Subtypes and IFNβ Influence Chronic Mucosal HIV-1 Pathogenesis

October 16, 2020

Kejun Guo, Guannan Shen, Jon Kibbie, Tania Gonzalez, Stephanie M Dillon, Harry A Smith, Emily H Cooper, Kerry Lavender, Kim J Hasenkrug, Kathrin Sutter, Ulf Dittmer, Miranda Kroehl, Katerina Kechris, Cara C Wilson and Mario L Santiago

The Type I Interferons (IFN-Is) are innate antiviral cytokines that include 12 different IFNα subtypes and IFNβ that signal through the IFN-I receptor (IFNAR), inducing hundreds of IFN-stimulated genes (ISGs) that comprise the 'interferome'.  Complete Abstract

The Crystal Structure of a Polerovirus Exoribonuclease-Resistant RNA Shows How Diverse Sequences are Integrated into a Conserved Fold

August 26, 2020

Anna-Lena Steckelberg, Quentin Vicens, David A. Costantino, Jay C. Nix and Jeffrey S. Kieft

Exoribonuclease-resistant RNAs (xrRNAs) are discrete elements that block the progression of 5′ to 3′ exoribonucleases using specifically folded RNA structures. A recently discovered class of xrRNA is widespread in several genera of plant-infecting viruses, within both...  Complete Abstract

JMJD5 couples with CDK9 to release the paused RNA polymerase II

August 18, 2020

Haolin Liu, Srinivas Ramachandran, Nova Fong, Tzu Phang, Schuyler Lee, Pirooz Parsa, Xinjian Liu, Laura Harmacek, Thomas Danhorn, Tengyao Song, Sangphil Oh, Qianqian Zhang, Zhongzhou Chen, Qian Zhang, Ting-Hui Tu, Carrie Happoldt, Brian O'Conner, Ralf Janknecht, Chuan-Yuan Li, Philippa Marrack, John Kappler, Sonia Leach and Gongyi Zhang

More than 30% of genes in higher eukaryotes are regulated by RNA polymerase II (Pol II) promoter proximal pausing. Pausing is released by the positive transcription elongation factor complex (P-TEFb).  Complete Abstract

Single-Cell RNA Sequencing of Childhood Ependymoma Reveals Neoplastic Cell Subpopulations that Impact Molecular Classification and Etiology

August 11, 2020

Austin E Gillen, Kent A Riemondy, Vladimir Amani, Andrea M Griesinger, Ahmed Gilani, Sujatha Venkataraman, Krishna Madhavan, Eric Prince, Bridget Sanford, Todd C Hankinson, Michael H Handler, Rajeev Vibhakar, Ken L Jones, Siddhartha Mitra, Jay R Hesselberth, Nicholas K Foreman and Andrew M Donson

Ependymoma (EPN) is a brain tumor commonly presenting in childhood that remains fatal in most children. Intra-tumoral cellular heterogeneity in bulk-tumor samples significantly confounds our understanding of EPN biology, impeding development of effective therapy. We, therefore, use single-cell RNA sequencing, histology, and deconvolution to catalog cellular heterogeneity of the major childhood EPN subgroups.  Complete Abstract

Simultaneous Measurement of Biochemical Phenotypes and Gene Expression in Single Cells

June 4, 2020

Amanda L Richer, Kent A Riemondy, Lakotah Hardie and Jay R Hesselberth

Methods to measure heterogeneity among cells are rapidly transforming our understanding of biology but are currently limited to molecular abundance measurements. We developed an approach to simultaneously measure biochemical activities and mRNA abundance in single cells... Complete Abstract

Gene–Diet Interactions: Dietary Rescue of Metabolic Defects in spen-Depleted Drosophila melanogaster

April 1, 2020

Claire M. Gillette, Kelsey E. Hazegh, Travis Nemkov, Davide Stefanoni, Angelo D’Alessandro, J. Matthew Taliaferro and Tania Reis

Obesity and its comorbidities are a growing health epidemic. Interactions between genetic background, the environment, and behavior (i.e., diet) greatly influence organismal energy balance. Previously, we described obesogenic mutations in the gene Split ends (Spen) in Drosophila melanogaster, and roles for Spen in fat storage and metabolic state.  Complete Abstract

An RNA Pseudoknot Stimulates HTLV-1 Pro-Pol Programmed -1 Ribosomal Frameshifting

January 26, 2020

E. Thulson, E. W. Hartwick, A. Cooper-Sansone, M. A. C. Williams, M. E. Soliman, L. K. Robinson, J. S. Kieft and K. D. Mouzakis

Programmed -1 ribosomal frameshifts (-1 PRFs) are commonly used by viruses to regulate their enzymatic and structural protein levels. Human T-cell leukemia virus type 1 (HTLV-1) is a carcinogenic retrovirus that uses two independent -1 PRFs to express viral enzymes critical to establishing new HTLV-1 infections.  Complete Abstract

Control of RNA Pol II Speed by PNUTS-PP1 and Spt5 Dephosphorylation Facilitates Termination by a "Sitting Duck Torpedo" Mechanism

November 4, 2019

M. A. Cortazar, R. M. Sheridan, B. Erickson, N. Fong, K. Glover-Cutter, K. Brannan and D. L. Bentley

Control of transcription speed, which influences many co-transcriptional processes, is poorly understood. We report that PNUTS-PP1 phosphatase is a negative regulator of RNA polymerase II (Pol II) elongation rate.  Complete Abstract

Coding Regions Affect mRNA Stability in Human Cells

September 19, 2019

A. Narula, J. Ellis, J. M. Taliaferro and O. S. Rissland

A new paradigm has emerged that coding regions can regulate mRNA stability in model organisms. Here, due to differences in cognate tRNA abundance, synonymous codons are translated at different speeds, and slow codons then stimulate mRNA decay.  Complete Abstract

How to Get Away with Nonsense: Mechanisms and Consequences of Escape from Nonsense-Mediated RNA Decay

July 31, 2019

M. C. Dyle, D. Kolakada, M. A. Cortazar and S. Jagannathan

Nonsense-mediated RNA decay (NMD) is an evolutionarily conserved RNA quality control process that serves both as a mechanism to eliminate aberrant transcripts carrying premature stop codons, and to regulate expression of some normal transcripts.  Complete Abstract

Classical and Emerging Techniques to Identify and Quantify Localized RNAs

May 3, 2019

J. M. Taliaferro

In essentially every cell, proteins are asymmetrically distributed according to their function. For many genes, this protein sorting problem is solved by transporting RNA molecules encoding the protein, rather than the protein itself, to the desired subcellular location.  Complete Abstract

Multiple Decay Events Target HAC1 mRNA During Splicing to Regulate the Unfolded Protein Response

March 16, 2019

P. D. Cherry, S. E. Peach and J. R. Hesselberth

In the unfolded protein response (UPR), stress in the endoplasmic reticulum (ER) activates a large transcriptional program to increase ER folding capacity. During the budding yeast UPR, Ire1 excises an intron from the HAC1 mRNA and the exon products of cleavage are ligated, and the translated protein induces hundreds of stress-response genes.  Complete Abstract