scholarly journals Erratum: Mapping Argonaute and conventional RNA-binding protein interactions with RNA at single-nucleotide resolution using HITS-CLIP and CIMS analysis

2016 ◽  
Vol 11 (3) ◽  
pp. 616-616 ◽  
Author(s):  
Michael J Moore ◽  
Chaolin Zhang ◽  
Emily Conn Gantman ◽  
Aldo Mele ◽  
Jennifer C Darnell ◽  
...  
Keyword(s):  
Protein Interactions ◽  
Rna Binding ◽  
Binding Protein ◽  
Rna Binding Protein ◽  
Single Nucleotide ◽  
Nucleotide Resolution ◽  
Single Nucleotide Resolution

scholarly journals Mapping Argonaute and conventional RNA-binding protein interactions with RNA at single-nucleotide resolution using HITS-CLIP and CIMS analysis

2014 ◽  
Vol 9 (2) ◽  
pp. 263-293 ◽  
Author(s):  
Michael J Moore ◽  
Chaolin Zhang ◽  
Emily Conn Gantman ◽  
Aldo Mele ◽  
Jennifer C Darnell ◽  
...  
Keyword(s):  
Protein Interactions ◽  
Rna Binding ◽  
Binding Protein ◽  
Rna Binding Protein ◽  
Single Nucleotide ◽  
Nucleotide Resolution ◽  
Single Nucleotide Resolution

scholarly journals Multivalent interactions with RNA drive RNA binding protein recruitment and dynamics in biomolecular condensates in Xenopus oocytes

2021 ◽  
Author(s):  
Sarah E Cabral ◽  
Kimberly Mowry
Keyword(s):  
Protein Interactions ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Binding Protein ◽  
Rna Binding Protein ◽  
Dependent Manner ◽  
Binding Domains ◽  
Multivalent Interactions

RNA localization and biomolecular condensate formation are key biological strategies for organizing the cytoplasm and generating cellular and developmental polarity. While enrichment of RNAs and RNA-binding proteins (RBPs) is a hallmark of both processes, the functional and structural roles of RNA-RNA and RNA-protein interactions within condensates remain unclear. Recent work from our laboratory has shown that RNAs required for germ layer patterning in Xenopus oocytes localize in novel biomolecular condensates, termed Localization bodies (L-bodies). L-bodies are composed of a non-dynamic RNA phase enmeshed in a more dynamic protein-containing phase. However, the interactions that drive the biophysical characteristics of L-bodies are not known. Here, we test the role of RNA-protein interactions using an L-body RNA-binding protein, PTBP3, which contains four RNA-binding domains (RBDs). We find that binding of RNA to PTB is required for both RNA and PTBP3 to be enriched in L-bodies in vivo. Importantly, while RNA binding to a single RBD is sufficient to drive PTBP3 localization to L-bodies, interactions between multiple RRMs and RNA tunes the dynamics of PTBP3 within L-bodies. In vitro, recombinant PTBP3 phase separates into non-dynamic structures in an RNA-dependent manner, supporting a role for RNA-protein interactions as a driver of both recruitment of components to L-bodies and the dynamics of the components after enrichment. Our results point to a model where RNA serves as a concentration-dependent, non-dynamic substructure and multivalent interactions with RNA are a key driver of protein dynamics.

scholarly journals Comparative analysis of LIN28-RNA binding sites identified at single nucleotide resolution

RNA Biology ◽  
2017 ◽  
Vol 14 (12) ◽  
pp. 1756-1765 ◽  
Author(s):  
Elizabeth Ransey ◽  
Anders Björkbom ◽  
Victor S. Lelyveld ◽  
Przemyslaw Biecek ◽  
Lorena Pantano ◽  
...  
Keyword(s):  
Comparative Analysis ◽  
Binding Sites ◽  
Rna Binding ◽  
Single Nucleotide ◽  
Nucleotide Resolution ◽  
Single Nucleotide Resolution ◽  
Rna Binding Sites

scholarly journals Substrate selectivity by the exonuclease Rrp6p

2019 ◽  
Vol 117 (2) ◽  
pp. 982-992 ◽  
Author(s):  
Armend Axhemi ◽  
Elizabeth V. Wasmuth ◽  
Christopher D. Lima ◽  
Eckhard Jankowsky
Keyword(s):  
Rna Binding ◽  
Large Range ◽  
Degradation Kinetics ◽  
Rna Decay ◽  
Substrate Selectivity ◽  
Single Nucleotide ◽  
Nucleotide Resolution ◽  
Single Nucleotide Resolution ◽  
In Cells

The exoribonuclease Rrp6p is critical for RNA decay in the nucleus. While Rrp6p acts on a large range of diverse substrates, it does not indiscriminately degrade all RNAs. How Rrp6p accomplishes this task is not understood. Here, we measure Rrp6p–RNA binding and degradation kinetics in vitro at single-nucleotide resolution and find an intrinsic substrate selectivity that enables Rrp6p to discriminate against specific RNAs. RNA length and the four 3′-terminal nucleotides contribute most to substrate selectivity and collectively enable Rrp6p to discriminate between different RNAs by several orders of magnitude. The most pronounced discrimination is seen against RNAs ending with CCA-3′. These RNAs correspond to 3′ termini of uncharged tRNAs, which are not targeted by Rrp6p in cells. The data show that in contrast to many other proteins that use substrate selectivity to preferentially interact with specific RNAs, Rrp6p utilizes its selectivity to discriminate against specific RNAs. This ability allows Rrp6p to target diverse substrates while avoiding a subset of RNAs.

scholarly journals Profiling the binding sites of RNA-binding protein by LACE-seq

2021 ◽  
Author(s):  
Ruibao Su ◽  
Di Wang ◽  
Changchang Cao ◽  
Yuanchao Xue
Keyword(s):  
Binding Sites ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Transcription Termination ◽  
Early Embryo ◽  
Linear Amplification ◽  
Single Nucleotide ◽  
Nucleotide Resolution ◽  
Single Nucleotide Resolution

Abstract RNA-binding proteins (RBPs) directly interact with various RNAs in living cells to regulate their processing, translation, and stability. Identifying the precise binding sites of RBPs is critical for appreciating their physiological or pathological roles in germline and early embryo development. Current methods typically need millions of cells to map RBP binding positions, which prevents us from appreciating the crucial role of RBPs in early development. Here, we present the LACE-seq method for unbiased mapping of RBP-binding sites at single-nucleotide resolution in fewer cells or even single oocytes. LACE-seq depends on RBP-mediated reverse transcription termination, and linear amplification of the cDNA ends for deep sequencing. To further promote its application, we describe a step-by-step protocol about how to construct a successful LACE-seq library.

scholarly journals A specific eIF4A paralog facilitates LARP1-mediated translation repression during mTORC1 inhibition

2021 ◽  
Author(s):  
Yuichi Shichino ◽  
Mari Mito ◽  
Kazuhiro Kashiwagi ◽  
Mari Takahashi ◽  
Takuhiro Ito ◽  
...  
Keyword(s):  
Translation Initiation ◽  
Protein Interactions ◽  
Rna Binding ◽  
Binding Protein ◽  
Rna Binding Protein ◽  
Ribosome Profiling ◽  
Translation Initiation Factor ◽  
Initiation Factor ◽  
Mtor Inhibition ◽  
Translation Repression

AbstractEukaryotic translation initiation factor (eIF) 4A — a DEAD-box RNA-binding protein — plays an essential role in translation initiation. Two mammalian eIF4A paralogs, eIF4A1 and eIF4A2, have been assumed to be redundant because of their high homology, and the difference in their functions has been poorly understood. Here, we show that eIF4A1, but not eIF4A2, enhances translational repression during the inhibition of mechanistic target of rapamycin complex 1 (mTORC1), an essential kinase complex controlling cell proliferation. RNA-immunoprecipitation sequencing (RIP-Seq) of the two eIF4A paralogs revealed that eIF4A1 preferentially binds to mRNAs containing terminal oligopyrimidine (TOP) motifs, whose translation is rapidly repressed upon mTOR inhibition. This biased interaction depends on a La-related RNA-binding protein, LARP1. Ribosome profiling revealed that the deletion of EIF4A1, but not EIF4A2, rendered the translation of TOP mRNAs resistant to mTOR inactivation. Moreover, eIF4A1 enhances the affinity between TOP mRNAs and LARP1 and thus ensures stronger translation repression upon mTORC1 inhibition. Our data show that the distinct protein interactions of these highly homologous translation factor paralogs shape protein synthesis during mTORC1 inhibition and provide a unique example of the repressive role of a universal translation activator.

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