scholarly journals Structural modularity of the XIST ribonucleoprotein complex

2020 ◽  
Vol 11 (1) ◽  
Author(s):  
Zhipeng Lu ◽  
Jimmy K. Guo ◽  
Yuning Wei ◽  
Diana R. Dou ◽  
Brian Zarnegar ◽  
...  
Keyword(s):  
Protein Complex ◽  
Protein Complexes ◽  
Effector Proteins ◽  
General Strategy ◽  
Sequence Motifs ◽  
Modular Architecture ◽  
Chimeric Rnas ◽  
Xist Rna ◽  
Evolutionarily Conserved ◽  
Associated Proteins

AbstractLong noncoding RNAs are thought to regulate gene expression by organizing protein complexes through unclear mechanisms. XIST controls the inactivation of an entire X chromosome in female placental mammals. Here we develop and integrate several orthogonal structure-interaction methods to demonstrate that XIST RNA-protein complex folds into an evolutionarily conserved modular architecture. Chimeric RNAs and clustered protein binding in fRIP and eCLIP experiments align with long-range RNA secondary structure, revealing discrete XIST domains that interact with distinct sets of effector proteins. CRISPR-Cas9-mediated permutation of the Xist A-repeat location shows that A-repeat serves as a nucleation center for multiple Xist-associated proteins and m6A modification. Thus modular architecture plays an essential role, in addition to sequence motifs, in determining the specificity of RBP binding and m6A modification. Together, this work builds a comprehensive structure-function model for the XIST RNA-protein complex, and suggests a general strategy for mechanistic studies of large ribonucleoprotein assemblies.

scholarly journals Structural modularity of the XIST ribonucleoprotein complex

2019 ◽  
Author(s):  
Zhipeng Lu ◽  
Jimmy K. Guo ◽  
Yuning Wei ◽  
Diana R. Dou ◽  
Brian Zarnegar ◽  
...  
Keyword(s):  
Protein Complex ◽  
Protein Complexes ◽  
Effector Proteins ◽  
General Strategy ◽  
Sequence Motifs ◽  
Modular Architecture ◽  
Chimeric Rnas ◽  
Xist Rna ◽  
Evolutionarily Conserved ◽  
Associated Proteins

SUMMARYLong noncoding RNAs are thought to regulate gene expression by organizing protein complexes through unclear mechanisms. XIST controls the inactivation of an entire X chromosome in female placental mammals. Here we develop and integrate several orthogonal structure-interaction methods to demonstrate that XIST RNA-protein complex folds into an evolutionarily conserved modular architecture. Chimeric RNAs and clustered protein binding in fRIP and eCLIP experiments align with long-range RNA secondary structure, revealing discrete XIST domains that interact with distinct sets of effector proteins. CRISPR-Cas9-mediated permutation of the Xist A-repeat location shows that A-repeat serves as a nucleation center for multiple Xist-associated proteins and m6A modification. Thus modular architecture plays an essential role, in addition to sequence motifs, in determining the specificity of RBP binding and m6A modification. Together, this work builds a comprehensive structure-function model for the XIST RNA-protein complex, and suggests a general strategy for mechanistic studies of large ribonucleoprotein assemblies.

scholarly journals The Sec34/Sec35p complex, a Ypt1p effector required for retrograde intra-Golgi trafficking, interacts with Golgi SNAREs and COPI vesicle coat proteins

2002 ◽  
Vol 157 (4) ◽  
pp. 631-643 ◽  
Author(s):  
Elena S. Suvorova ◽  
Rainer Duden ◽  
Vladimir V. Lupashin
Keyword(s):  
Protein Complex ◽  
Protein Complexes ◽  
Protein Sorting ◽  
Golgi Vesicle ◽  
Secretory Proteins ◽  
Coat Proteins ◽  
Rab Protein ◽  
Golgi Membranes ◽  
Evolutionarily Conserved ◽  
Copi Vesicle

The Sec34/35 complex was identified as one of the evolutionarily conserved protein complexes that regulates a cis-Golgi step in intracellular vesicular transport. We have identified three new proteins that associate with Sec35p and Sec34p in yeast cytosol. Mutations in these Sec34/35 complex subunits result in defects in basic Golgi functions, including glycosylation of secretory proteins, protein sorting, and retention of Golgi resident proteins. Furthermore, the Sec34/35 complex interacts genetically and physically with the Rab protein Ypt1p, intra-Golgi SNARE molecules, as well as with Golgi vesicle coat complex COPI. We propose that the Sec34/35 protein complex acts as a tether that connects cis-Golgi membranes and COPI-coated, retrogradely targeted intra-Golgi vesicles.

scholarly journals Identification of a Novel RAMA/RON3 Rhoptry Protein Complex in Plasmodium falciparum Merozoites

2021 ◽  
Vol 10 ◽  
Author(s):  
Daisuke Ito ◽  
Jun-Hu Chen ◽  
Eizo Takashima ◽  
Tomoyuki Hasegawa ◽  
Hitoshi Otsuki ◽  
...  
Keyword(s):  
Plasmodium Falciparum ◽  
Monoclonal Antibodies ◽  
Molecular Mass ◽  
Protein Complex ◽  
Protein Complexes ◽  
Parasite Development ◽  
Erythrocyte Invasion ◽  
Liquid Chromatography Tandem Mass ◽  
Associated Proteins ◽  
Rhoptry Protein

Malaria causes a half a million deaths annually. The parasite intraerythrocytic lifecycle in the human bloodstream is the major cause of morbidity and mortality. Apical organelles of merozoite stage parasites are involved in the invasion of erythrocytes. A limited number of apical organellar proteins have been identified and characterized for their roles during erythrocyte invasion or subsequent intraerythrocytic parasite development. To expand the repertoire of identified apical organellar proteins we generated a panel of monoclonal antibodies against Plasmodium falciparum schizont-rich parasites and screened the antibodies using immunofluorescence assays. Out of 164 hybridoma lines, 12 clones produced monoclonal antibodies yielding punctate immunofluorescence staining patterns in individual merozoites in late schizonts, suggesting recognition of merozoite apical organelles. Five of the monoclonal antibodies were used to immuno-affinity purify their target antigens and these antigens were identified by liquid chromatography-tandem mass spectrometry (LC-MS/MS). Two known apical organelle protein complexes were identified, the high-molecular mass rhoptry protein complex (PfRhopH1/Clags, PfRhopH2, and PfRhopH3) and the low-molecular mass rhoptry protein complex (rhoptry-associated proteins complex, PfRAP1, and PfRAP2). A novel complex was additionally identified by immunoprecipitation, composed of rhoptry-associated membrane antigen (PfRAMA) and rhoptry neck protein 3 (PfRON3) of P. falciparum. We further identified a region spanning amino acids Q221-E481 within the PfRAMA that may associate with PfRON3 in immature schizonts. Further investigation will be required as to whether PfRAMA and PfRON3 interact directly or indirectly.

Protein Interaction Domains and Post-Translational Modifications: Structural Features and Drug Discovery Applications

2020 ◽  
Vol 27 (37) ◽  
pp. 6306-6355 ◽  
Author(s):  
Marian Vincenzi ◽  
Flavia Anna Mercurio ◽  
Marilisa Leone
Keyword(s):  
Drug Discovery ◽  
Protein Interaction ◽  
Protein Interactions ◽  
Structural Information ◽  
Protein Complexes ◽  
Structural Features ◽  
Protein Protein Interactions ◽  
Modular Architecture ◽  
Post Translational Modifications ◽  
Interaction Domains

Background:: Many pathways regarding healthy cells and/or linked to diseases onset and progression depend on large assemblies including multi-protein complexes. Protein-protein interactions may occur through a vast array of modules known as protein interaction domains (PIDs). Objective:: This review concerns with PIDs recognizing post-translationally modified peptide sequences and intends to provide the scientific community with state of art knowledge on their 3D structures, binding topologies and potential applications in the drug discovery field. Method:: Several databases, such as the Pfam (Protein family), the SMART (Simple Modular Architecture Research Tool) and the PDB (Protein Data Bank), were searched to look for different domain families and gain structural information on protein complexes in which particular PIDs are involved. Recent literature on PIDs and related drug discovery campaigns was retrieved through Pubmed and analyzed. Results and Conclusion:: PIDs are rather versatile as concerning their binding preferences. Many of them recognize specifically only determined amino acid stretches with post-translational modifications, a few others are able to interact with several post-translationally modified sequences or with unmodified ones. Many PIDs can be linked to different diseases including cancer. The tremendous amount of available structural data led to the structure-based design of several molecules targeting protein-protein interactions mediated by PIDs, including peptides, peptidomimetics and small compounds. More studies are needed to fully role out, among different families, PIDs that can be considered reliable therapeutic targets, however, attacking PIDs rather than catalytic domains of a particular protein may represent a route to obtain selective inhibitors.

scholarly journals Humanizing the yeast origin recognition complex

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Clare S. K. Lee ◽  
Ming Fung Cheung ◽  
Jinsen Li ◽  
Yongqian Zhao ◽  
Wai Hei Lam ◽  
...  
Keyword(s):  
Amino Acid ◽  
Protein Complex ◽  
Origin Recognition Complex ◽  
Specific Binding ◽  
Life Cycles ◽  
Amino Acid Insertion ◽  
Evolutionarily Conserved ◽  
Transcriptional Start Sites ◽  
Eukaryote Genome ◽  
Origin Recognition

AbstractThe Origin Recognition Complex (ORC) is an evolutionarily conserved six-subunit protein complex that binds specific sites at many locations to coordinately replicate the entire eukaryote genome. Though highly conserved in structure, ORC’s selectivity for replication origins has diverged tremendously between yeasts and humans to adapt to vastly different life cycles. In this work, we demonstrate that the selectivity determinant of ORC for DNA binding lies in a 19-amino acid insertion helix in the Orc4 subunit, which is present in yeast but absent in human. Removal of this motif from Orc4 transforms the yeast ORC, which selects origins based on base-specific binding at defined locations, into one whose selectivity is dictated by chromatin landscape and afforded with plasticity, as reported for human. Notably, the altered yeast ORC has acquired an affinity for regions near transcriptional start sites (TSSs), which the human ORC also favors.

scholarly journals Binding of an X-Specific Condensin Correlates with a Reduction in Active Histone Modifications at Gene Regulatory Elements

Genetics ◽  
2019 ◽  
Vol 212 (3) ◽  
pp. 729-742 ◽  
Author(s):  
Lena Annika Street ◽  
Ana Karina Morao ◽  
Lara Heermans Winterkorn ◽  
Chen-Yu Jiao ◽  
Sarah Elizabeth Albritton ◽  
...  
Keyword(s):  
Gene Expression ◽  
Histone Modifications ◽  
Chromatin Immunoprecipitation ◽  
Protein Complexes ◽  
Regulatory Elements ◽  
Evolutionarily Conserved ◽  
Chromatin Immunoprecipitation Sequencing ◽  
Gene Regulatory Elements ◽  
Gene Regulatory ◽  
Regulatory Sites

Condensins are evolutionarily conserved protein complexes that are required for chromosome segregation during cell division and genome organization during interphase. In Caenorhabditis elegans, a specialized condensin, which forms the core of the dosage compensation complex (DCC), binds to and represses X chromosome transcription. Here, we analyzed DCC localization and the effect of DCC depletion on histone modifications, transcription factor binding, and gene expression using chromatin immunoprecipitation sequencing and mRNA sequencing. Across the X, the DCC accumulates at accessible gene regulatory sites in active chromatin and not heterochromatin. The DCC is required for reducing the levels of activating histone modifications, including H3K4me3 and H3K27ac, but not repressive modification H3K9me3. In X-to-autosome fusion chromosomes, DCC spreading into the autosomal sequences locally reduces gene expression, thus establishing a direct link between DCC binding and repression. Together, our results indicate that DCC-mediated transcription repression is associated with a reduction in the activity of X chromosomal gene regulatory elements.

scholarly journals Atypical Protein Kinase C Is Involved in the Evolutionarily Conserved Par Protein Complex and Plays a Critical Role in Establishing Epithelia-Specific Junctional Structures

2001 ◽  
Vol 152 (6) ◽  
pp. 1183-1196 ◽  
Author(s):  
Atsushi Suzuki ◽  
Tomoyuki Yamanaka ◽  
Tomonori Hirose ◽  
Naoyuki Manabe ◽  
Keiko Mizuno ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
Cell Polarity ◽  
Protein Complex ◽  
Mdck Cells ◽  
Critical Role ◽  
Dominant Negative ◽  
Dominant Negative Mutant ◽  
Surface Polarity ◽  
C Elegans ◽  
Evolutionarily Conserved

We have previously shown that during early Caenorhabditis elegans embryogenesis PKC-3, a C. elegans atypical PKC (aPKC), plays critical roles in the establishment of cell polarity required for subsequent asymmetric cleavage by interacting with PAR-3 [Tabuse, Y., Y. Izumi, F. Piano, K.J. Kemphues, J. Miwa, and S. Ohno. 1998. Development (Camb.). 125:3607–3614]. Together with the fact that aPKC and a mammalian PAR-3 homologue, aPKC-specific interacting protein (ASIP), colocalize at the tight junctions of polarized epithelial cells (Izumi, Y., H. Hirose, Y. Tamai, S.-I. Hirai, Y. Nagashima, T. Fujimoto, Y. Tabuse, K.J. Kemphues, and S. Ohno. 1998. J. Cell Biol. 143:95–106), this suggests a ubiquitous role for aPKC in establishing cell polarity in multicellular organisms. Here, we show that the overexpression of a dominant-negative mutant of aPKC (aPKCkn) in MDCK II cells causes mislocalization of ASIP/PAR-3. Immunocytochemical analyses, as well as measurements of paracellular diffusion of ions or nonionic solutes, demonstrate that the biogenesis of the tight junction structure itself is severely affected in aPKCkn-expressing cells. Furthermore, these cells show increased interdomain diffusion of fluorescent lipid and disruption of the polarized distribution of Na+,K+-ATPase, suggesting that epithelial cell surface polarity is severely impaired in these cells. On the other hand, we also found that aPKC associates not only with ASIP/PAR-3, but also with a mammalian homologue of C. elegans PAR-6 (mPAR-6), and thereby mediates the formation of an aPKC-ASIP/PAR-3–PAR-6 ternary complex that localizes to the apical junctional region of MDCK cells. These results indicate that aPKC is involved in the evolutionarily conserved PAR protein complex, and plays critical roles in the development of the junctional structures and apico-basal polarization of mammalian epithelial cells.

scholarly journals DrosophilaTorsin Protein Regulates Motor Control and Stress Sensitivity and Forms a Complex with Fragile-X Mental Retardation Protein

2016 ◽  
Vol 2016 ◽  
pp. 1-14 ◽  
Author(s):  
Phuong Nguyen ◽  
Jong Bok Seo ◽  
Hyo-Min Ahn ◽  
Young Ho Koh
Keyword(s):  
Mental Retardation ◽  
Protein Complexes ◽  
Fragile X ◽  
Stress Sensitivity ◽  
Altered Expression ◽  
Fragile X Mental Retardation ◽  
Evolutionarily Conserved ◽  
Mental Retardation Protein ◽  
Synaptic Boutons

We investigated unknownin vivofunctions of Torsin by usingDrosophilaas a model. Downregulation ofDrosophilaTorsin (DTor) by DTor-specific inhibitory double-stranded RNA (RNAi) induced abnormal locomotor behavior and increased susceptibility to H2O2. In addition, altered expression of DTor significantly increased the numbers of synaptic boutons. One important biochemical consequence of DTor-RNAi expression in fly brains was upregulation of alcohol dehydrogenase (ADH). Altered expression of ADH has also been reported inDrosophilaFragile-X mental retardation protein (DFMRP) mutant flies. Interestingly, expression of DFMRP was altered in DTor mutant flies, and DTor and DFMRP were present in the same protein complexes. In addition, DTor and DFMRP immunoreactivities were partially colocalized in several cellular organelles in larval muscles. Furthermore, there were no significant differences between synaptic morphologies ofdfmrpnull mutants anddfmrpmutants expressing DTor-RNAi. Taken together, our evidences suggested that DTor and DFMRP might be present in the same signaling pathway regulating synaptic plasticity. In addition, we also found that human Torsin1A and human FMRP were present in the same protein complexes, suggesting that this phenomenon is evolutionarily conserved.

scholarly journals TAP (NXF1) Belongs to a Multigene Family of Putative RNA Export Factors with a Conserved Modular Architecture

2000 ◽  
Vol 20 (23) ◽  
pp. 8996-9008 ◽  
Author(s):  
Andrea Herold ◽  
Mikita Suyama ◽  
João P. Rodrigues ◽  
Isabelle C. Braun ◽  
Ulrike Kutay ◽  
...  
Keyword(s):  
Drosophila Melanogaster ◽  
Rna Binding ◽  
Mrna Export ◽  
Modular Architecture ◽  
Export Activity ◽  
Evolutionarily Conserved ◽  
Rna Export ◽  
Higher Eukaryotes ◽  
Leucine Rich Repeats ◽  
Rna Export Factors

ABSTRACT Vertebrate TAP (also called NXF1) and its yeast orthologue, Mex67p, have been implicated in the export of mRNAs from the nucleus. The TAP protein includes a noncanonical RNP-type RNA binding domain, four leucine-rich repeats, an NTF2-like domain that allows heterodimerization with p15 (also called NXT1), and a ubiquitin-associated domain that mediates the interaction with nucleoporins. Here we show that TAP belongs to an evolutionarily conserved family of proteins that has more than one member in higher eukaryotes. Not only the overall domain organization but also residues important for p15 and nucleoporin interaction are conserved in most family members. We characterize two of four human TAP homologues and show that one of them, NXF2, binds RNA, localizes to the nuclear envelope, and exhibits RNA export activity. NXF3, which does not bind RNA or localize to the nuclear rim, has no RNA export activity. Database searches revealed that although only one p15(nxt) gene is present in the Drosophila melanogaster and Caenorhabditis elegans genomes, there is at least one additional p15 homologue (p15-2 [also called NXT2]) encoded by the human genome. Both human p15 homologues bind TAP, NXF2, and NXF3. Together, our results indicate that the TAP-p15 mRNA export pathway has diversified in higher eukaryotes compared to yeast, perhaps reflecting a greater substrate complexity.

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