scholarly journals An Interaction between Two RNA Binding Proteins, Nab2 and Pub1, Links mRNA Processing/Export and mRNA Stability

2007 ◽  
Vol 27 (18) ◽  
pp. 6569-6579 ◽  
Author(s):  
Luciano H. Apponi ◽  
Seth M. Kelly ◽  
Michelle T. Harreman ◽  
Alexander N. Lehner ◽  
Anita H. Corbett ◽  
...  
Keyword(s):  
Mrna Stability ◽  
Binding Proteins ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Binding Protein ◽  
Tail Length ◽  
Functional Link ◽  
Mrna Binding Protein ◽  
Mrna Binding

ABSTRACT mRNA stability is modulated by elements in the mRNA transcript and their cognate RNA binding proteins. Poly(U) binding protein 1 (Pub1) is a cytoplasmic Saccharomyces cerevisiae mRNA binding protein that stabilizes transcripts containing AU-rich elements (AREs) or stabilizer elements (STEs). In a yeast two-hybrid screen, we identified nuclear poly(A) binding protein 2 (Nab2) as being a Pub1-interacting protein. Nab2 is an essential nucleocytoplasmic shuttling mRNA binding protein that regulates poly(A) tail length and mRNA export. The interaction between Pub1 and Nab2 was confirmed by copurification and in vitro binding assays. The interaction is mediated by the Nab2 zinc finger domain. Analysis of the functional link between these proteins reveals that Nab2, like Pub1, can modulate the stability of specific mRNA transcripts. The half-life of the RPS16B transcript, an ARE-like sequence-containing Pub1 target, is decreased in both nab2-1 and nab2-67 mutants. In contrast, GCN4, an STE-containing Pub1 target, is not affected. Similar results were obtained for other ARE- and STE-containing Pub1 target transcripts. Further analysis reveals that the ARE-like sequence is necessary for Nab2-mediated transcript stabilization. These results suggest that Nab2 functions together with Pub1 to modulate mRNA stability and strengthen a model where nuclear events are coupled to the control of mRNA turnover in the cytoplasm.

Role Of Insulin Like Growth Factor mRNA Binding Protein-3 (IGF2BP3) In Mixed Lineage Leukemia (MLL) Positive B-Cell Lymphomas

Blood ◽  
2013 ◽  
Vol 122 (21) ◽  
pp. 3816-3816
Author(s):  
Jayanth Kumar Palanichamy ◽  
Tiffany Tran ◽  
Jorge Contreras ◽  
Thilini R Fernando ◽  
Dinesh S. Rao
Keyword(s):  
Gene Expression ◽  
Growth Factor ◽  
B Cell ◽  
Cell Line ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Binding Protein ◽  
Insulin Like Growth Factor ◽  
Mrna Binding Protein ◽  
Mrna Binding

Abstract Oncogenic transformation of early B-cell progenitors leads to the human disease of B-acute lymphoblastic leukemia or B-ALL, which affects both children and adults. Among the different subtypes of B-ALL, defined by particular cytogenetic anomalies, there are two which are difficult to treat and have a dismal prognosis. These are B-ALL with chromosomal translocation t (9; 22)/BCR-ABL and MLL gene rearrangements, which show distinctive gene expression profiles. Gene expression is now known to be significantly regulated by post-transcriptional mechanisms. These involve RNA binding proteins and microRNAs. Deregulation of microRNAs as well as RNA binding protein expression is associated with numerous cancers. Here, we hypothesized that RNA binding proteins may be important in regulating gene expression in MLL rearranged leukemias. To examine this hypothesis, we undertook a microarray study examining the expression of both protein-coding and non-coding genes in B-ALL, including MLL translocations. A total of 44 samples were used for the microarray. Supervised class prediction was carried out using the R library of prediction analysis for microarrays (PAM). One of the most significantly differentially expressed genes was Insulin Like Growth Factor mRNA Binding Protein-3 (IGF2BP3). The expression of IGF2BP3 was highest in the MLL rearranged B-ALL group. IGF2BP3 is an oncofetal protein known to be highly expressed in a number of epithelial malignancies such as glioblastomas. IGF2BP3 has been known to bind to the 5’-UTR and stabilize mRNAs like CD44 the expression of which correlates with epithelial tumors metastasis. IGF2BP3 has been shown to bind to the Insulin like Growth Factor-2 (IGF-2) mRNA and enhance translation in glioblastomas. We confirmed the expression of IGF2BP3 and CD44 in these 44 tumor samples and 90 other B-cell lymphoma samples by RT-qPCR. This corroborated with our previous data showing that the expression of both these genes is significantly higher in the group with MLL translocations. In the MLL rearranged leukemias, there was a significant correlation between the expression of CD44 and IGF2BP3. Interestingly however, there was no significant difference in the expression of IGF2 mRNA between these different subsets, indicating either that IGF2BP3 might be acting on IGF-2 mRNA at the translational level or that IGF-2 regulation may be cell-type specific. To evaluate whether IGF2BP3 affects the growth of B-ALL cells, we used NALM6, a B-ALL cell line which expresses IGF2BP3. We generated microRNA-155 formatted siRNAs against human IGF2BP3 and subcloned them into pHAGE6 based lentiviral vectors. Our preliminary data demonstrates that these vectors are capable of knocking down IGF2BP3 in the NALM6 cell line. In addition, cells with knockdown showed a dramatic decrease in their growth rates, as measured by the MTS assay. The IGF-2 paracrine signaling system is thought to be important in the maintenance of HSCs as well as in lymphocyte development. We separated different precursors of B-cells (Hardy fractions) from murine bone marrow using FACS and measured the mRNA expression of CD44 and IGF2BP3 in these different subsets. The expression of both these genes correlated well with each other and showed a dynamic expression pattern with the highest expression seen in the Hardy Fraction C (late pro-B cells). This indicates that the IGF2BP3/CD44 axis might play a role in regulating normal B-cell development and this may be dysregulated in MLL-translocated B-ALL. To examine whether IGF2BP3 overexpression causes leukemia, we cloned the murine and human IGF2BP3 coding regions in a murine retroviral expression vector, MIG (MSCV-IGF2BP3-IRES-GFP). Retroviral packaging was done using 293T cell line, virus was collected and used to infect 7Oz/3, a murine pre-B ALL cell line. Western blot and qPCR confirmed overexpression of IGF2BP3. We have infected bone marrow cells from CD 45.2 positive wild type donor mice with the virus and transferred them into irradiated CD 45.1 recipient mice. We have confirmed engraftment in these mice using flowcytometry for CD 45.1/2 and are presently following the mice for the development of leukemia. In summary, IGF2BP3 is dysregulated in MLL-rearranged leukemia, and its knockdown can cause decreased growth rates in B-ALL cell lines. The current study explores whether IGF2BP3 is oncogenic and the mechanisms of action of IGF2BP3 in B-cell development and neoplasia. Disclosures: No relevant conflicts of interest to declare.

scholarly journals The 36-kilodalton embryonic-type cytoplasmic polyadenylation element-binding protein in Xenopus laevis is ElrA, a member of the ELAV family of RNA-binding proteins.

1997 ◽  
Vol 17 (11) ◽  
pp. 6402-6409 ◽  
Author(s):  
L Wu ◽  
P J Good ◽  
J D Richter
Keyword(s):  
Xenopus Laevis ◽  
Binding Proteins ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Binding Protein ◽  
Dominant Negative ◽  
Cytoplasmic Polyadenylation ◽  
Element Binding Protein

The translational activation of several maternal mRNAs in Xenopus laevis is dependent on cytoplasmic poly(A) elongation. Messages harboring the UUUUUAU-type cytoplasmic polyadenylation element (CPE) in their 3' untranslated regions (UTRs) undergo polyadenylation and translation during oocyte maturation. This CPE is bound by the protein CPEB, which is essential for polyadenylation. mRNAs that have the poly(U)12-27 embryonic-type CPE (eCPE) in their 3' UTRs undergo polyadenylation and translation during the early cleavage and blastula stages. A 36-kDa eCPE-binding protein in oocytes and embryos has been identified by UV cross-linking. We now report that this 36-kDa protein is ElrA, a member of the ELAV family of RNA-binding proteins. The proteins are identical in size, antibody directed against ElrA immunoprecipitates the 36-kDa protein, and the two proteins have the same RNA binding specificity in vitro. C12 and activin receptor mRNAs, both of which contain eCPEs, are detected in immunoprecipitated ElrA-mRNP complexes from eggs and embryos. In addition, this in vivo interaction requires the eCPE. Although a number of experiments failed to define a role for ElrA in cytoplasmic polyadenylation, the expression of a dominant negative ElrA protein in embryos results in an exogastrulation phenotype. The possible functions of ElrA in gastrulation are discussed.

scholarly journals The RNA-binding protein PTBP1 promotes ATPase-dependent dissociation of the RNA helicase UPF1 to protect transcripts from nonsense-mediated mRNA decay

2020 ◽  
Vol 295 (33) ◽  
pp. 11613-11625 ◽  
Author(s):  
Sarah E. Fritz ◽  
Soumya Ranganathan ◽  
Clara D. Wang ◽  
J. Robert Hogg
Keyword(s):  
Binding Proteins ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Binding Protein ◽  
Target Selection ◽  
Rna Helicase ◽  
Nonsense Mediated Decay ◽  
Polypyrimidine Tract Binding Protein ◽  
Regulatory Loop

The sequence-specific RNA-binding proteins PTBP1 (polypyrimidine tract–binding protein 1) and HNRNP L (heterogeneous nuclear ribonucleoprotein L) protect mRNAs from nonsense-mediated decay (NMD) by preventing the UPF1 RNA helicase from associating with potential decay targets. Here, by analyzing in vitro helicase activity, dissociation of UPF1 from purified mRNPs, and transcriptome-wide UPF1 RNA binding, we present the mechanistic basis for inhibition of NMD by PTBP1. Unlike mechanisms of RNA stabilization that depend on direct competition for binding sites among protective RNA-binding proteins and decay factors, PTBP1 promotes displacement of UPF1 already bound to potential substrates. Our results show that PTBP1 directly exploits the tendency of UPF1 to release RNA upon ATP binding and hydrolysis. We further find that UPF1 sensitivity to PTBP1 is coordinated by a regulatory loop in domain 1B of UPF1. We propose that the UPF1 regulatory loop and protective proteins control kinetic proofreading of potential NMD substrates, presenting a new model for RNA helicase regulation and target selection in the NMD pathway.

scholarly journals The Roles of Insulin-Like Growth Factor 2 mRNA-Binding Protein 2 in Cancer and Cancer Stem Cells

2018 ◽  
Vol 2018 ◽  
pp. 1-15 ◽  
Author(s):  
Junguo Cao ◽  
Qingchun Mu ◽  
Haiyan Huang
Keyword(s):  
Stem Cells ◽  
Growth Factor ◽  
Cancer Stem Cells ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Binding Protein ◽  
Insulin Like Growth Factor ◽  
Wide Range ◽  
Mrna Binding Protein ◽  
Mrna Binding

RNA-binding proteins (RBPs) mediate the localization, stability, and translation of the target transcripts and fine-tune the physiological functions of the proteins encoded. The insulin-like growth factor (IGF) 2 mRNA-binding protein (IGF2BP, IMP) family comprises three RBPs, IGF2BP1, IGF2BP2, and IGF2BP3, capable of associating with IGF2 and other transcripts and mediating their processing. IGF2BP2 represents the least understood member of this family of RBPs; however, it has been reported to participate in a wide range of physiological processes, such as embryonic development, neuronal differentiation, and metabolism. Its dysregulation is associated with insulin resistance, diabetes, and carcinogenesis and may potentially be a powerful biomarker and candidate target for relevant diseases. This review summarizes the structural features, regulation, and functions of IGF2BP2 and their association with cancer and cancer stem cells.

scholarly journals Single-molecule imaging reveals dynamic biphasic partition of RNA-binding proteins in stress granules

2018 ◽  
Vol 217 (4) ◽  
pp. 1303-1318 ◽  
Author(s):  
Benedikt Niewidok ◽  
Maxim Igaev ◽  
Abel Pereira da Graca ◽  
Andre Strassner ◽  
Christine Lenzen ◽  
...  
Keyword(s):  
Liquid Phase ◽  
Single Molecule ◽  
Mammalian Cells ◽  
Binding Proteins ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Liquid Droplet ◽  
Binding Protein ◽  
Stress Granules

Stress granules (SGs) are cytosolic, nonmembranous RNA–protein complexes. In vitro experiments suggested that they are formed by liquid–liquid phase separation; however, their properties in mammalian cells remain unclear. We analyzed the distribution and dynamics of two paradigmatic RNA-binding proteins (RBPs), Ras GTPase-activating protein SH3-domain–binding protein (G3BP1) and insulin-like growth factor II mRNA-binding protein 1 (IMP1), with single-molecule resolution in living neuronal cells. Both RBPs exhibited different exchange kinetics between SGs. Within SGs, single-molecule localization microscopy revealed distributed hotspots of immobilized G3BP1 and IMP1 that reflect the presence of relatively immobile nanometer-sized nanocores. We demonstrate alternating binding in nanocores and anomalous diffusion in the liquid phase with similar characteristics for both RBPs. Reduction of low-complexity regions in G3BP1 resulted in less detectable mobile molecules in the liquid phase without change in binding in nanocores. The data provide direct support for liquid droplet behavior of SGs in living cells and reveal transient binding of RBPs in nanocores. Our study uncovers a surprising disconnect between SG partitioning and internal diffusion and interactions of RBPs.

scholarly journals Evolutionarily Conserved Polyadenosine RNA Binding Protein Nab2 Cooperates with Splicing Machinery To Regulate the Fate of Pre-mRNA

2016 ◽  
Vol 36 (21) ◽  
pp. 2697-2714 ◽  
Author(s):  
Sharon Soucek ◽  
Yi Zeng ◽  
Deepti L. Bellur ◽  
Megan Bergkessel ◽  
Kevin J. Morris ◽  
...  
Keyword(s):  
Binding Proteins ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Binding Protein ◽  
Rna Binding Protein ◽  
Tail Length ◽  
Mrna Processing ◽  
Evolutionarily Conserved ◽  
Mrna Maturation

Numerous RNA binding proteins are deposited onto an mRNA transcript to modulate posttranscriptional processing events ensuring proper mRNA maturation. Defining the interplay between RNA binding proteins that couple mRNA biogenesis events is crucial for understanding how gene expression is regulated. To explore how RNA binding proteins control mRNA processing, we investigated a role for the evolutionarily conserved polyadenosine RNA binding protein, Nab2, in mRNA maturation within the nucleus. This study reveals thatnab2mutant cells accumulate intron-containing pre-mRNAin vivo. We extend this analysis to identify genetic interactions between mutant alleles ofnab2and genes encoding a splicing factor,MUD2, and RNA exosome,RRP6, within vivoconsequences of altered pre-mRNA splicing and poly(A) tail length control. As further evidence linking Nab2 proteins to splicing, an unbiased proteomic analysis of vertebrate Nab2, ZC3H14, identifies physical interactions with numerous components of the spliceosome. We validated the interaction between ZC3H14 and U2AF2/U2AF65. Taking all the findings into consideration, we present a model where Nab2/ZC3H14 interacts with spliceosome components to allow proper coupling of splicing with subsequent mRNA processing steps contributing to a kinetic proofreading step that allows properly processed mRNA to exit the nucleus and escape Rrp6-dependent degradation.

scholarly journals The Interplay of RNA Binding Proteins, Oxidative Stress and Mitochondrial Dysfunction in ALS

Antioxidants ◽  
2021 ◽  
Vol 10 (4) ◽  
pp. 552
Author(s):  
Jasmine Harley ◽  
Benjamin E. Clarke ◽  
Rickie Patani
Keyword(s):  
Oxidative Stress ◽  
Gene Expression ◽  
Mitochondrial Dysfunction ◽  
Binding Proteins ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Binding Protein ◽  
Rna Binding Protein ◽  
Therapeutic Strategies ◽  
Lateral Sclerosis

RNA binding proteins fulfil a wide number of roles in gene expression. Multiple mechanisms of RNA binding protein dysregulation have been implicated in the pathomechanisms of several neurodegenerative diseases including amyotrophic lateral sclerosis (ALS). Oxidative stress and mitochondrial dysfunction also play important roles in these diseases. In this review, we highlight the mechanistic interplay between RNA binding protein dysregulation, oxidative stress and mitochondrial dysfunction in ALS. We also discuss different potential therapeutic strategies targeting these pathways.

scholarly journals Identification of mRNAs Associated with αCP2-Containing RNP Complexes

2003 ◽  
Vol 23 (19) ◽  
pp. 7055-7067 ◽  
Author(s):  
Shelly A. Waggoner ◽  
Stephen A. Liebhaber
Keyword(s):  
Binding Proteins ◽  
Rna Binding ◽  
Cell Function ◽  
Rna Binding Proteins ◽  
Translation Control ◽  
Viral Mrnas ◽  
Posttranscriptional Gene Regulation ◽  
Direct Binding

ABSTRACT Posttranscriptional controls in higher eukaryotes are central to cell differentiation and developmental programs. These controls reflect sequence-specific interactions of mRNAs with one or more RNA binding proteins. The α-globin poly(C) binding proteins (αCPs) comprise a highly abundant subset of K homology (KH) domain RNA binding proteins and have a characteristic preference for binding single-stranded C-rich motifs. αCPs have been implicated in translation control and stabilization of multiple cellular and viral mRNAs. To explore the full contribution of αCPs to cell function, we have identified a set of mRNAs that associate in vivo with the major αCP2 isoforms. One hundred sixty mRNA species were consistently identified in three independent analyses of αCP2-RNP complexes immunopurified from a human hematopoietic cell line (K562). These mRNAs could be grouped into subsets encoding cytoskeletal components, transcription factors, proto-oncogenes, and cell signaling factors. Two mRNAs were linked to ceroid lipofuscinosis, indicating a potential role for αCP2 in this infantile neurodegenerative disease. Surprisingly, αCP2 mRNA itself was represented in αCP2-RNP complexes, suggesting autoregulatory control of αCP2 expression. In vitro analyses of representative target mRNAs confirmed direct binding of αCP2 within their 3′ untranslated regions. These data expand the list of mRNAs that associate with αCP2 in vivo and establish a foundation for modeling its role in coordinating pathways of posttranscriptional gene regulation.

scholarly journals Matrin3: Disorder and ALS Pathogenesis

2022 ◽  
Vol 8 ◽  
Author(s):  
Ahmed Salem ◽  
Carter J. Wilson ◽  
Benjamin S. Rutledge ◽  
Allison Dilliott ◽  
Sali Farhan ◽  
...  
Keyword(s):  
Nuclear Export ◽  
Binding Proteins ◽  
Rna Binding ◽  
Nuclear Dna ◽  
Neurodegenerative Disorder ◽  
Rna Binding Proteins ◽  
Binding Protein ◽  
Intrinsically Disordered ◽  
Intrinsically Disordered Regions ◽  
Disordered Regions

Amyotrophic lateral sclerosis (ALS) is a neurodegenerative disorder characterized by the degeneration of both upper and lower motor neurons in the brain and spinal cord. ALS is associated with protein misfolding and inclusion formation involving RNA-binding proteins, including TAR DNA-binding protein (TDP-43) and fused in sarcoma (FUS). The 125-kDa Matrin3 is a highly conserved nuclear DNA/RNA-binding protein that is implicated in many cellular processes, including binding and stabilizing mRNA, regulating mRNA nuclear export, modulating alternative splicing, and managing chromosomal distribution. Mutations in MATR3, the gene encoding Matrin3, have been identified as causal in familial ALS (fALS). Matrin3 lacks a prion-like domain that characterizes many other ALS-associated RNA-binding proteins, including TDP-43 and FUS, however, our bioinformatics analyses and preliminary studies document that Matrin3 contains long intrinsically disordered regions that may facilitate promiscuous interactions with many proteins and may contribute to its misfolding. In addition, these disordered regions in Matrin3 undergo numerous post-translational modifications, including phosphorylation, ubiquitination and acetylation that modulate the function and misfolding of the protein. Here we discuss the disordered nature of Matrin3 and review the factors that may promote its misfolding and aggregation, two elements that might explain its role in ALS pathogenesis.

scholarly journals SSMART: Sequence-structure motif identification for RNA-binding proteins

2018 ◽  
Author(s):  
Alina Munteanu ◽  
Neelanjan Mukherjee ◽  
Uwe Ohler
Keyword(s):  
Binding Sites ◽  
Binding Proteins ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Sequence Motif ◽  
Primary Sequence ◽  
Sequence Structure ◽  
Rna Targets

AbstractMotivationRNA-binding proteins (RBPs) regulate every aspect of RNA metabolism and function. There are hundreds of RBPs encoded in the eukaryotic genomes, and each recognize its RNA targets through a specific mixture of RNA sequence and structure properties. For most RBPs, however, only a primary sequence motif has been determined, while the structure of the binding sites is uncharacterized.ResultsWe developed SSMART, an RNA motif finder that simultaneously models the primary sequence and the structural properties of the RNA targets sites. The sequence-structure motifs are represented as consensus strings over a degenerate alphabet, extending the IUPAC codes for nucleotides to account for secondary structure preferences. Evaluation on synthetic data showed that SSMART is able to recover both sequence and structure motifs implanted into 3‘UTR-like sequences, for various degrees of structured/unstructured binding sites. In addition, we successfully used SSMART on high-throughput in vivo and in vitro data, showing that we not only recover the known sequence motif, but also gain insight into the structural preferences of the RBP.AvailabilitySSMART is freely available at https://ohlerlab.mdc-berlin.de/software/SSMART_137/[email protected]

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