Telomerase Biogenesis: RNA Processing, Trafficking, and Protein Interactions

This study reports the HMGB1 interactomes in prostate and ovary cancer cells lines. Affinity purification coupled to mass spectrometry confirmed that the HMGB1 nuclear interactome is involved in HMGB1 known functions such as maintenance of chromatin stability and regulation of transcription, and also in not as yet reported processes such as mRNA and rRNA processing. We have identified an interaction between HMGB1 and the NuRD complex and validated this by yeast-two-hybrid, confirming that the RBBP7 subunit directly interacts with HMGB1. In addition, we describe for the first time an interaction between two HMGB1 interacting complexes, the septin and THOC complexes, as well as an interaction of these two complexes with Rab11. Analysis of Pan-Cancer Atlas public data indicated that several genes encoding HMGB1-interacting proteins identified in this study are dysregulated in tumours from patients diagnosed with ovary and prostate carcinomas. In PC-3 cells, silencing of HMGB1 leads to downregulation of the expression of key regulators of ribosome biogenesis and RNA processing, namely BOP1, RSS1, UBF1, KRR1 and LYAR. Upregulation of these genes in prostate adenocarcinomas is correlated with worse prognosis, reinforcing their functional significance in cancer progression.

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Nuclear RNA-protein interactions and messenger RNA processing.

The Journal of Cell Biology ◽

10.1083/jcb.97.5.1321 ◽

1983 ◽

Vol 97 (5) ◽

pp. 1321-1326 ◽

Cited By ~ 32

Author(s):

T Pederson

Keyword(s):

Detailed Analysis ◽

Recent Work ◽

Protein Interactions ◽

Rna Processing ◽

Messenger Rna ◽

Mammalian Cells ◽

General Hypothesis ◽

Ribonucleoprotein Particles ◽

Nuclear Rna ◽

Nuclear Ribonucleoprotein

Eucaryotic messenger RNA precursors are processed in nuclear ribonucleoprotein particles (hnRNP). Here recent work on the structure of hnRNP is reviewed, with emphasis on function. Detailed analysis of a specific case, the altered assembly of hnRNP in heat-shocked Drosophila and mammalian cells, leads to a general hypothesis linking hnRNP structure and messenger RNA processing.

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Targeting RNA Binding Proteins Involved in Neurodegeneration

CrossRef Listing of Deleted DOIs ◽

10.1177/1087057113497256 ◽

2013 ◽

Vol 18 (9) ◽

pp. 967-983 ◽

Cited By ~ 15

Author(s):

Maurizio Romano ◽

Emanuele Buratti

Keyword(s):

Protein Interactions ◽

Rna Processing ◽

Noncoding Rna ◽

Rna Binding ◽

Rna Binding Proteins ◽

Correct Identification ◽

Protein Protein Interactions ◽

Aggregation Processes ◽

Cellular Pathways ◽

Screening Approaches

Dysfunctions at the level of RNA processing have recently been shown to play a fundamental role in the pathogenesis of many neurodegenerative diseases. Several proteins responsible for these dysfunctions (TDP-43, FUS/TLS, and hnRNP A/Bs) belong to the nuclear class of heterogeneous ribonucleoproteins (hnRNPs) that predominantly function as general regulators of both coding and noncoding RNA metabolism. The discovery of the importance of these factors in mediating neuronal death has represented a major paradigmatic shift in our understanding of neurodegenerative processes. As a result, these discoveries have also opened the way toward novel biomolecular screening approaches in our search for therapeutic options. One of the major hurdles in this search is represented by the correct identification of the most promising targets to be prioritized. These may include aberrant aggregation processes, protein-protein interactions, RNA-protein interactions, or specific cellular pathways altered by disease. In this review, we discuss these four major options together with their various advantages and drawbacks.

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Targeting translation activity at the ribosome interface with UV-active small molecules

10.1101/436311 ◽

2018 ◽

Author(s):

Divya T. Kandala ◽

Alessia Del Piano ◽

Luca Minati ◽

Massimiliano Clamer

Keyword(s):

Protein Synthesis ◽

Small Molecules ◽

Protein Interactions ◽

Rna Processing ◽

Translational Control ◽

Control Mechanisms ◽

Translation Efficiency ◽

Catalytic Core ◽

Genome Wide

ABSTRACTPuromycin is a well-known antibiotic that is used to study the mechanism of protein synthesis and to monitor translation efficiency due to its incorporation into nascent peptide chains. However, puromycin effects outside the ribo-some catalytic core remain unexplored. Here, we developed two puromycin analogues (3PB and 3PC) that can efficiently interact with several proteins involved in translation, ribosome function and RNA processing. We biochemically characterized the binding of these analogues and globally mapped the direct small molecule-protein interactions in living cells using clickable and photoreactive puromycin-like probes in combination with in-depth mass spectrometry. We identified a list of proteins that interact with ribosomes during translation (e.g. eEF1A, ENO1 and GRP78) and we addressed possible uses of the probes to sense the activity of protein synthesis and to capture associated RNA. By coupling genome-wide RNA sequencing methods with these molecules, the characterization of unexplored translational control mechanisms will be feasible.

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POST-TRANSCRIPTIONAL GENE REGULATION: RNA-PROTEIN INTERACTIONS, RNA PROCESSING, MRNA STABILITY AND LOCALIZATION

Biocomputing 2009 ◽

10.1142/9789812836939_0052 ◽

2008 ◽

Author(s):

BENJAMIN BLENCOWE ◽

STEVEN BRENNER ◽

TIMOTHY HUGHES ◽

QUAID MORRIS

Keyword(s):

Gene Regulation ◽

Protein Interactions ◽

Rna Processing ◽

Mrna Stability ◽

Transcriptional Gene Regulation

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Affinity proteomic dissection of the human nuclear cap-binding complex interactome

Nucleic Acids Research ◽

10.1093/nar/gkaa743 ◽

2020 ◽

Vol 48 (18) ◽

pp. 10456-10469 ◽

Cited By ~ 1

Author(s):

Yuhui Dou ◽

Svetlana Kalmykova ◽

Maria Pashkova ◽

Mehrnoosh Oghbaie ◽

Hua Jiang ◽

...

Keyword(s):

Rna Polymerase Ii ◽

Protein Interactions ◽

Rna Processing ◽

Biological Pathways ◽

Experimental Conditions ◽

Affinity Capture ◽

Macromolecular Assemblies ◽

Cap Binding Complex ◽

Rna Exosome ◽

Incomplete Models

Abstract A 5′,7-methylguanosine cap is a quintessential feature of RNA polymerase II-transcribed RNAs, and a textbook aspect of co-transcriptional RNA processing. The cap is bound by the cap-binding complex (CBC), canonically consisting of nuclear cap-binding proteins 1 and 2 (NCBP1/2). Interest in the CBC has recently renewed due to its participation in RNA-fate decisions via interactions with RNA productive factors as well as with adapters of the degradative RNA exosome. A novel cap-binding protein, NCBP3, was recently proposed to form an alternative CBC together with NCBP1, and to interact with the canonical CBC along with the protein SRRT. The theme of post-transcriptional RNA fate, and how it relates to co-transcriptional ribonucleoprotein assembly, is abundant with complicated, ambiguous, and likely incomplete models. In an effort to clarify the compositions of NCBP1-, 2- and 3-related macromolecular assemblies, we have applied an affinity capture-based interactome screen where the experimental design and data processing have been modified to quantitatively identify interactome differences between targets under a range of experimental conditions. This study generated a comprehensive view of NCBP-protein interactions in the ribonucleoprotein context and demonstrates the potential of our approach to benefit the interpretation of complex biological pathways.

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The 40-kilodalton to autoantigen associates with nucleotides 21 to 64 of human mitochondrial RNA processing/7-2 RNA in vitro

Molecular and Cellular Biology ◽

10.1128/mcb.11.10.5266-5274.1991 ◽

1991 ◽

Vol 11 (10) ◽

pp. 5266-5274

Author(s):

Y Yuan ◽

E Tan ◽

R Reddy

Keyword(s):

Binding Site ◽

Protein Interactions ◽

Sequence Homology ◽

Rna Processing ◽

Tertiary Structure ◽

Rnase P ◽

Cell Extract ◽

Mitochondrial Rna ◽

Antigen Binding Site

A 40-kDa To antigen recognized by sera from some patients with autoimmune diseases is an integral component of both human RNase P and mitochondrial RNA processing (MRP) RNase. Human MRP and RNase P RNAs, synthesized in vitro, readily associate with the To antigen present in the HeLa cell extract. Using this in vitro reconstitution system, the binding site of the To antigen is localized to a 44-nucleotide-long sequence corresponding to nucleotides 21 to 64 of the human MRP RNA. UV cross-linking experiments showed that the To antigen binds directly to MRP RNA and to RNase P (H1) RNA through RNA-protein interactions. Although the MRP RNA and RNAse P (H1) RNA show sequence homology in four conserved blocks (H. A. Gold, J. N. Topper, D. A. Clayton, and J. Craft, Science 245:1377-1380, 1989), the To antigen-binding site in MRP RNA does not show any obvious primary sequence homology with H1 RNA. These data suggest that the To antigen binds to a conserved and presumably a common secondary or tertiary structure in human MRP and RNase P RNAs.

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Novel functional insights revealed by distinct protein-protein interactions of the residual SWI/SNF complex in SMARCA4-deficient small cell carcinoma of the ovary, hypercalcemic type

10.1101/794776 ◽

2019 ◽

Cited By ~ 1

Author(s):

Elizabeth A. Raupach ◽

Krystine Garcia-Mansfield ◽

Ritin Sharma ◽

Apurva M. Hegde ◽

Victoria David-Dirgo ◽

...

Keyword(s):

Cell Lines ◽

Chromatin Remodeling ◽

Protein Interactions ◽

Rna Processing ◽

Small Cell ◽

Wild Type ◽

Protein Protein Interactions ◽

Functional Consequences ◽

Carcinoma Of The Ovary ◽

Inactivating Mutations

AbstractChromatin remodeling plays a critical role in tumor suppression as demonstrated by 20% of human cancers bearing inactivating mutations in SWI/SNF chromatin remodeling complex members. Mutations in different SWI/SNF subunits drive a variety of adult and pediatric tumor types, including non-small cell lung cancers, rhabdoid tumors, medulloblastomas, and ovarian cancers. Small cell carcinoma of the ovary hypercalcemic type (SCCOHT) is an aggressive subtype of ovarian cancer occurring in young women. Nearly all (>98%) SCCOHTs have inactivating mutations in SMARCA4, which encodes 1 of 2 mutually exclusive catalytic subunits of the SWI/SNF complex. Less than half of SCCOHT patients survive 5 years despite aggressive surgery and multimodal chemotherapy. Empirical support for effective SCCOHT treatments is scarce, in part because of the poor understanding of SCCOHT tumorigenesis. To gain insight into the functional consequences of SWI/SNF subunit loss, we defined SWI/SNF composition and its protein-protein interactions (PPIs) by immunoprecipitation and mass spectrometry (IP-MS) of SWI/SNF subunits in 3 SCCOHT cell lines. Comparing these results to a cell line containing a wild-type SWI/SNF complex, the interaction of most canonical core SWI/SNF subunits was observed in all SCCOHT cell lines at a lower abundance. The SCCOHT SWI/SNF also lacked ATPase module subunits and showed a drastic reduction in PBAF-specific subunit interactions. The wild-type and SCCOHT SWI/SNF subunits immunoprecipitated a shared set of 26 proteins, including core SWI/SNF subunits and RNA processing proteins. We observed 131 proteins exclusively interacting with the wild-type SWI/SNF complex including isoform-specific SWI/SNF subunits, members of the NuRD complex, and members of the MLL3/4 complex. We observed 60 PPIs exclusive to the SCCOHT residual SWI/SNF shared in at least 2 of the 3 SCCOHT cell lines, including many proteins involved in RNA processing. Differential interactions with the residual SWI/SNF complex in SCCOHT may further elucidate altered functional consequences of SMARCA4 mutations in these tumors as well as identify synthetic lethal targets that translate to other SWI/SNF-deficient tumors.

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Protein Interaction Screen on Peptide Matrix (PRISMA) reveals interaction footprints and the PTM-dependent interactome of intrinsically disordered C/EBPβ

10.1101/238709 ◽

2017 ◽

Author(s):

Gunnar Dittmar ◽

Daniel Perez Hernandez ◽

Elisabeth Kowenz-Leutz ◽

Marieluise Kirchner ◽

Günther Kahlert ◽

...

Keyword(s):

Cell Differentiation ◽

Cell Fate ◽

Protein Interactions ◽

Rna Processing ◽

Intrinsically Disordered Proteins ◽

Disordered Proteins ◽

List Type ◽

Molecular Feature ◽

Intrinsically Disordered ◽

Cell Extracts

SummaryCCAAT enhancer binding protein beta (C/EBPβ) is a pioneer transcription factor that specifies cell differentiation. C/EBPβ is intrinsically unstructured, a molecular feature common to many proteins involved in signal processing and epigenetics. The structure of C/EBPβ differs depending on alternative translation start site usage and multiple post-translational modifications (PTM). Mutation of distinct PTM sites in C/EBPβ alters designated protein interactions and cell differentiation, suggesting a C/EBPβ PTM indexing code determines epigenetic outcomes. Herein, we systematically explored the interactome of C/EBPβ using an array of spot-synthesised C/EBPβ-derived linear tiling peptides with and without PTM, combined with mass spectrometric proteomic analysis of protein interactions. We identified interaction footprints of ~1300 proteins in nuclear cell extracts, many with chromatin modifying, remodelling and RNA processing functions. The results suggest C/EBPβ acts as a multi-tasking molecular switchboard, integrating signal-dependent modifications and structural plasticity to orchestrate interactions with numerous protein complexes directing cell fate and function.HighlightsPeptide array based interaction proteomics map SLiM and PTM dependent C/EBPβ interactomeNovel links between C/EBPβ, RNA processing, transcription elongation, MLL, NuRD were revealedC/EBPβ structure organizes modular hub function for gene regulatory machineryPRISMA is suitable to resolve protein interactions and networks based on intrinsically disordered proteins

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The 40-kilodalton to autoantigen associates with nucleotides 21 to 64 of human mitochondrial RNA processing/7-2 RNA in vitro.

Molecular and Cellular Biology ◽

10.1128/mcb.11.10.5266 ◽

1991 ◽

Vol 11 (10) ◽

pp. 5266-5274 ◽

Cited By ~ 34

Author(s):

Y Yuan ◽

E Tan ◽

R Reddy

Keyword(s):

Binding Site ◽

Protein Interactions ◽

Sequence Homology ◽

Rna Processing ◽

Tertiary Structure ◽

Rnase P ◽

Cell Extract ◽

Mitochondrial Rna ◽

Antigen Binding Site

A 40-kDa To antigen recognized by sera from some patients with autoimmune diseases is an integral component of both human RNase P and mitochondrial RNA processing (MRP) RNase. Human MRP and RNase P RNAs, synthesized in vitro, readily associate with the To antigen present in the HeLa cell extract. Using this in vitro reconstitution system, the binding site of the To antigen is localized to a 44-nucleotide-long sequence corresponding to nucleotides 21 to 64 of the human MRP RNA. UV cross-linking experiments showed that the To antigen binds directly to MRP RNA and to RNase P (H1) RNA through RNA-protein interactions. Although the MRP RNA and RNAse P (H1) RNA show sequence homology in four conserved blocks (H. A. Gold, J. N. Topper, D. A. Clayton, and J. Craft, Science 245:1377-1380, 1989), the To antigen-binding site in MRP RNA does not show any obvious primary sequence homology with H1 RNA. These data suggest that the To antigen binds to a conserved and presumably a common secondary or tertiary structure in human MRP and RNase P RNAs.

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