The Experts below are selected from a list of 270 Experts worldwide ranked by ideXlab platform

Joel P. Mackay - One of the best experts on this subject based on the ideXlab platform.

  • the zinc fingers of yy1 bind Single Stranded RNA with low sequence specificity
    Nucleic Acids Research, 2016
    Co-Authors: Dorothy C C Wai, Manar Shihab, Jason Low, Joel P. Mackay
    Abstract:

    Classical zinc fingers (ZFs) are traditionally considered to act as sequence-specific DNA-binding domains. More recently, classical ZFs have been recognised as potential RNA-binding modules, raising the intriguing possibility that classical-ZF transcription factors are involved in post-transcriptional gene regulation via direct RNA binding. To date, however, only one classical ZF-RNA complex, that involving TFIIIA, has been structurally characterised. Yin Yang-1 (YY1) is a multi-functional transcription factor involved in many regulatory processes, and binds DNA via four classical ZFs. Recent evidence suggests that YY1 also interacts with RNA, but the molecular nature of the interaction remains unknown. In the present work, we directly assess the ability of YY1 to bind RNA using in vitro assays. Systematic Evolution of Ligands by EXponential enrichment (SELEX) was used to identify preferred RNA sequences bound by the YY1 ZFs from a randomised library over multiple rounds of selection. However, a strong motif was not consistently recovered, suggesting that the RNA sequence selectivity of these domains is modest. YY1 ZF residues involved in binding to Single-Stranded RNA were identified by NMR spectroscopy and found to be largely distinct from the set of residues involved in DNA binding, suggesting that interactions between YY1 and ssRNA constitute a separate mode of nucleic acid binding. Our data are consistent with recent reports that YY1 can bind to RNA in a low-specificity, yet physiologically relevant manner.

  • Engineering Specificity Changes on a RanBP2 Zinc Finger that Binds Single-Stranded RNA†
    Angewandte Chemie (International ed. in English), 2014
    Co-Authors: Marylène Vandevenne, Mitchell R. O’connell, Stephanie Helder, Nicholas E. Shepherd, Jacqueline M. Matthews, Ann H. Kwan, David J. Segal, Joel P. Mackay
    Abstract:

    The realization that gene transcription is much more pervasive than previously thought and that many diverse RNA species exist in simple as well as complex organisms has triggered efforts to develop functionalized RNA-binding proteins (RBPs) that have the ability to probe and manipulate RNA function. Previously, we showed that the RanBP2-type zinc finger (ZF) domain is a good candidate for an addressable Single-Stranded-RNA (ssRNA) binding domain that can recognize ssRNA in a modular and specific manner. In the present study, we successfully engineered a sequence specificity change onto this ZF scaffold by using a combinatorial approach based on phage display. This work constitutes a foundation from which a set of RanBP2 ZFs might be developed that is able to recognize any given RNA sequence.

Junjie Zhang - One of the best experts on this subject based on the ideXlab platform.

  • Cryo-EM reveals infection steps of Single-Stranded RNA bacteriophages.
    Progress in biophysics and molecular biology, 2020
    Co-Authors: Karl V. Gorzelnik, Junjie Zhang
    Abstract:

    Abstract Single-Stranded RNA bacteriophages (ssRNA phages) are small spherical RNA viruses that infect bacteria with retractile pili. The Single positive-sense genomic RNA of ssRNA phages, which is protected by a capsid shell, is delivered into the host via the retraction of the host pili. Structures involved in ssRNA phage infection cycle are essential for understanding the underlying mechanisms that can be used to engineer them for therapeutic applications. This review summarizes the recent breakthroughs in high-resolution structural studies of two ssRNA phages, MS2 and Qβ, and their interaction with the host, E. coli, by cryo-electron microscopy (cryo-EM). These studies revealed new cryo-EM structures, which provide insights into how MS2 and Qβ package the RNA, lyse E. coli, and adsorb to the receptor F-pili, responsible for conjugation. Methodologies described here can be expanded to study other ssRNA phages that target pathogenic bacteria.

  • Structural basis for the adsorption of a Single-Stranded RNA bacteriophage
    Nature Communications, 2019
    Co-Authors: Ran Meng, Mengqiu Jiang, Zhicheng Cui, Jeng-yih Chang, Kailu Yang, Joanita Jakana, Zhao Wang, Junjie Zhang
    Abstract:

    Single-Stranded RNA bacteriophages use a Single maturation protein (Mat) to attach to a retractile pilus of the bacterial host. Here, the authors report the structures of the MS2 phage bound to the host receptor F-pili and define the orientations of Mat relative to the cell and emanating F-pili, providing new insights into the F-like type IV secretion systems. Single-Stranded RNA bacteriophages (ssRNA phages) infect Gram-negative bacteria via a Single maturation protein (Mat), which attaches to a retractile pilus of the host. Here we present structures of the ssRNA phage MS2 in complex with the Escherichia coli F-pilus, showing a network of hydrophobic and electrostatic interactions at the Mat-pilus interface. Moreover, binding of the pilus induces slight orientational variations of the Mat relative to the rest of the phage capsid, priming the Mat-connected genomic RNA (gRNA) for its release from the virions. The exposed tip of the attached Mat points opposite to the direction of the pilus retraction, which may facilitate the translocation of the gRNA from the capsid into the host cytosol. In addition, our structures determine the orientation of the assembled F-pilin subunits relative to the cell envelope, providing insights into the F-like type IV secretion systems.

  • Structural basis for the adsorption of a Single-Stranded RNA bacteriophage
    Nature communications, 2019
    Co-Authors: Ran Meng, Mengqiu Jiang, Zhicheng Cui, Jeng-yih Chang, Kailu Yang, Joanita Jakana, Zhao Wang, Junjie Zhang
    Abstract:

    Single-Stranded RNA bacteriophages (ssRNA phages) infect Gram-negative bacteria via a Single maturation protein (Mat), which attaches to a retractile pilus of the host. Here we present structures of the ssRNA phage MS2 in complex with the Escherichia coli F-pilus, showing a network of hydrophobic and electrostatic interactions at the Mat-pilus interface. Moreover, binding of the pilus induces slight orientational variations of the Mat relative to the rest of the phage capsid, priming the Mat-connected genomic RNA (gRNA) for its release from the virions. The exposed tip of the attached Mat points opposite to the direction of the pilus retraction, which may facilitate the translocation of the gRNA from the capsid into the host cytosol. In addition, our structures determine the orientation of the assembled F-pilin subunits relative to the cell envelope, providing insights into the F-like type IV secretion systems.

Janggi Choi - One of the best experts on this subject based on the ideXlab platform.

  • lower and upper stem Single Stranded RNA junctions together determine the drosha cleavage site
    Proceedings of the National Academy of Sciences of the United States of America, 2013
    Co-Authors: Yonggan Wu, Janggi Choi, Haoquan Wu
    Abstract:

    Microprocessor [Drosha–DGCR8 (DiGeorge syndrome critical region gene 8) complex] processing of primary microRNA (pri-miRNA) is the critical first step in miRNA biogenesis, but how the Drosha cleavage site is determined has been unclear. Previous models proposed that the Drosha–DGCR8 complex measures either ∼22 nt from the upper stem–Single-Stranded RNA (ssRNA, terminal loop) junction or ∼11 nt from the lower stem–ssRNA junction to determine the cleavage site. Here, using miRNA-offset RNAs to determine the Drosha cleavage site, we show that the Microprocessor measures the distances from both the lower and upper stem–ssRNA junctions to determine the cleavage site in human cells, and optimal distances from both structures are critical to the precision of Drosha processing. If the distances are not optimal, Drosha tends to cleave at multiple sites, which can, in turn, generate multiple 5′ isomiRs. Thus, our results also reveal a mechanism of 5′ isomiR generation.

  • Lower and upper stem–Single-Stranded RNA junctions together determine the Drosha cleavage site
    Proceedings of the National Academy of Sciences of the United States of America, 2013
    Co-Authors: Janggi Choi
    Abstract:

    Microprocessor [Drosha–DGCR8 (DiGeorge syndrome critical region gene 8) complex] processing of primary microRNA (pri-miRNA) is the critical first step in miRNA biogenesis, but how the Drosha cleavage site is determined has been unclear. Previous models proposed that the Drosha–DGCR8 complex measures either ∼22 nt from the upper stem–Single-Stranded RNA (ssRNA, terminal loop) junction or ∼11 nt from the lower stem–ssRNA junction to determine the cleavage site. Here, using miRNA-offset RNAs to determine the Drosha cleavage site, we show that the Microprocessor measures the distances from both the lower and upper stem–ssRNA junctions to determine the cleavage site in human cells, and optimal distances from both structures are critical to the precision of Drosha processing. If the distances are not optimal, Drosha tends to cleave at multiple sites, which can, in turn, generate multiple 5′ isomiRs. Thus, our results also reveal a mechanism of 5′ isomiR generation.

Haoquan Wu - One of the best experts on this subject based on the ideXlab platform.

  • lower and upper stem Single Stranded RNA junctions together determine the drosha cleavage site
    Proceedings of the National Academy of Sciences of the United States of America, 2013
    Co-Authors: Yonggan Wu, Janggi Choi, Haoquan Wu
    Abstract:

    Microprocessor [Drosha–DGCR8 (DiGeorge syndrome critical region gene 8) complex] processing of primary microRNA (pri-miRNA) is the critical first step in miRNA biogenesis, but how the Drosha cleavage site is determined has been unclear. Previous models proposed that the Drosha–DGCR8 complex measures either ∼22 nt from the upper stem–Single-Stranded RNA (ssRNA, terminal loop) junction or ∼11 nt from the lower stem–ssRNA junction to determine the cleavage site. Here, using miRNA-offset RNAs to determine the Drosha cleavage site, we show that the Microprocessor measures the distances from both the lower and upper stem–ssRNA junctions to determine the cleavage site in human cells, and optimal distances from both structures are critical to the precision of Drosha processing. If the distances are not optimal, Drosha tends to cleave at multiple sites, which can, in turn, generate multiple 5′ isomiRs. Thus, our results also reveal a mechanism of 5′ isomiR generation.

Martin Zacharias - One of the best experts on this subject based on the ideXlab platform.

  • fragment based modelling of Single Stranded RNA bound to RNA recognition motif containing proteins
    Nucleic Acids Research, 2016
    Co-Authors: Isaure Chauvot De Beauchene, Sjoerd J De Vries, Martin Zacharias
    Abstract:

    Protein-RNA complexes are important for many biological processes. However, structural modeling of such complexes is hampered by the high flexibility of RNA. Particularly challenging is the docking of Single-Stranded RNA (ssRNA). We have developed a fragment-based approach to model the structure of ssRNA bound to a protein, based on only the protein structure, the RNA sequence and conserved contacts. The conformational diversity of each RNA fragment is sampled by an exhaustive library of trinucleotides extracted from all known experimental protein-RNA complexes. The method was applied to ssRNA with up to 12 nucleotides which bind to dimers of the RNA recognition motifs (RRMs), a highly abundant eukaryotic RNA-binding domain. The fragment based docking allows a precise de novo atomic modeling of protein-bound ssRNA chains. On a benchmark of seven experimental ssRNA-RRM complexes, near-native models (with a mean heavy-atom deviation of <3 A from experiment) were generated for six out of seven bound RNA chains, and even more precise models (deviation < 2 A) were obtained for five out of seven cases, a significant improvement compared to the state of the art. The method is not restricted to RRMs but was also successfully applied to Pumilio RNA binding proteins.

  • Fragment-based modelling of Single Stranded RNA bound to RNA recognition motif containing proteins
    Nucleic Acids Research, 2016
    Co-Authors: Isaure Chauvot De Beauchêne, Sjoerd De Vries, Martin Zacharias
    Abstract:

    Protein-RNA complexes are important for many biological processes. However, structural modeling of such complexes is hampered by the high flexibility of RNA. Particularly challenging is the docking of Single-Stranded RNA (ssRNA). We have developed a fragment-based approach to model the structure of ssRNA bound to a protein, based on only the protein structure, the RNA sequence and conserved contacts. The conformational diversity of each RNA fragment is sampled by an exhaustive library of trinucleotides extracted from all known experimental protein-RNA complexes. The method was applied to ssRNA with up to 12 nucleotides which bind to dimers of the RNA recognition motifs (RRMs), a highly abundant eukaryotic RNA-binding domain. The fragment based docking allows a precise de novo atomic modeling of protein-bound ssRNA chains. On a benchmark of seven experimental ssRNA-RRM complexes, near-native models (with a mean heavy-atom deviation of