Browse
Submit Data
Databases
API
Help

Dataset Information

0 Views

0 Connections

0 Citations

0 Reanalyses

0 Downloads

Omics score: 0

Mechanism of ?-actin mRNA Recognition by ZBP1.

ABSTRACT: Zipcode binding protein 1 (ZBP1) is an oncofetal RNA-binding protein that mediates the transport and local translation of ?-actin mRNA by the KH3-KH4 di-domain, which is essential for neuronal development. The high-resolution structures of KH3-KH4 with their respective target sequences show that KH4 recognizes a non-canonical GGA sequence via an enlarged and dynamic hydrophobic groove, whereas KH3 binding to a core CA sequence occurs with low specificity. A data-informed kinetic simulation of the two-step binding reaction reveals that the overall reaction is driven by the second binding event and that the moderate affinities of the individual interactions favor RNA looping. Furthermore, the concentration of ZBP1, but not of the target RNA, modulates the interaction, which explains the functional significance of enhanced ZBP1 expression during embryonic development.

SUBMITTER: Nicastro G

PROVIDER: S-EPMC5300891 | biostudies-literature | 2017 Jan

REPOSITORIES: biostudies-literature

ACCESS DATA

Json Xml

Publications

Mechanism of β-actin mRNA Recognition by ZBP1.

Nicastro Giuseppe G Candel Adela M AM Uhl Michael M Oregioni Alain A Hollingworth David D Backofen Rolf R Martin Stephen R SR Ramos Andres A

Cell reports 20170101 5

Zipcode binding protein 1 (ZBP1) is an oncofetal RNA-binding protein that mediates the transport and local translation of β-actin mRNA by the KH3-KH4 di-domain, which is essential for neuronal development. The high-resolution structures of KH3-KH4 with their respective target sequences show that KH4 recognizes a non-canonical GGA sequence via an enlarged and dynamic hydrophobic groove, whereas KH3 binding to a core CA sequence occurs with low specificity. A data-informed kinetic simulation of th ...[more]

PMID: 28147274

Similar Datasets

ZBP1 recognition of beta-actin zipcode induces RNA looping.

Project description:ZBP1 (zipcode-binding protein 1) was originally discovered as a trans-acting factor for the "zipcode" in the 3' untranslated region (UTR) of the beta-actin mRNA that is important for its localization and translational regulation. Subsequently, ZBP1 has been found to be a multifunctional regulator of RNA metabolism that controls aspects of localization, stability, and translation for many mRNAs. To reveal how ZBP1 recognizes its RNA targets, we biochemically characterized the interaction between ZBP1 and the beta-actin zipcode. The third and fourth KH (hnRNP K homology) domains of ZBP1 specifically recognize a bipartite RNA element located within the first 28 nucleotides of the zipcode. The spacing between the RNA sequences is consistent with the structure of IMP1 KH34, the human ortholog of ZBP1, that we solved by X-ray crystallography. The tandem KH domains are arranged in an intramolecular anti-parallel pseudodimer conformation with the canonical RNA-binding surfaces at opposite ends of the molecule. This orientation of the KH domains requires that the RNA backbone must undergo an approximately 180 degrees change in direction in order for both KH domains to contact the RNA simultaneously. The RNA looping induced by ZBP1 binding provides a mechanism for specific recognition and may facilitate the assembly of post-transcriptional regulatory complexes by remodeling the bound transcript.

| S-EPMC2807350 | biostudies-literature

ZBP2 facilitates binding of ZBP1 to beta-actin mRNA during transcription.

Project description:Cytoplasmic mRNA localization regulates gene expression by spatially restricting protein translation. Recent evidence has shown that nuclear proteins (such as hnRNPs) are required to form mRNPs capable of cytoplasmic localization. ZBP1 and ZBP2, two hnRNP K homology domain-containing proteins, were previously identified by their binding to the zipcode, the sequence element necessary and sufficient for beta-actin mRNA localization. ZBP1 colocalizes with nascent beta-actin mRNA in the nucleus but is predominantly a cytoplasmic protein. ZBP2, in contrast, is predominantly nuclear. We hypothesized that the two proteins cooperate to localize beta-actin mRNA and sought to address where and how this might occur. We demonstrate that ZBP2, a homologue of the splicing factor KSRP, binds initially to nascent beta-actin transcripts and facilitates the subsequent binding of the shuttling ZBP1. ZBP1 then associates with the RNA throughout the nuclear export and cytoplasmic localization process.

| S-EPMC2169170 | biostudies-literature

RACK1 is a ribosome scaffold protein for ?-actin mRNA/ZBP1 complex.

Project description:In neurons, specific mRNAs are transported in a translationally repressed manner along dendrites or axons by transport ribonucleic-protein complexes called RNA granules. ZBP1 is one RNA binding protein present in transport RNPs, where it transports and represses the translation of cotransported mRNAs, including ?-actin mRNA. The release of ?-actin mRNA from ZBP1 and its subsequent translation depends on the phosphorylation of ZBP1 by Src kinase, but little is known about how this process is regulated. Here we demonstrate that the ribosomal-associated protein RACK1, another substrate of Src, binds the ?-actin mRNA/ZBP1 complex on ribosomes and contributes to the release of ?-actin mRNA from ZBP1 and to its translation. We identify the Src binding and phosphorylation site Y246 on RACK1 as the critical site for the binding to the ?-actin mRNA/ZBP1 complex. Based on these results we propose RACK1 as a ribosomal scaffold protein for specific mRNA-RBP complexes to tightly regulate the translation of specific mRNAs.

| S-EPMC3327689 | biostudies-literature

Real-time visualization of ZBP1 association with beta-actin mRNA during transcription and localization.

Project description:BACKGROUND:mRNA localization in somatic cells is an important mechanism for gene expression regulation. In fibroblasts, the protein ZBP1 associates with the sequence that localizes beta-actin mRNA to the leading edge of fibroblasts, augmenting motility. beta-actin mRNA localizes in a cytoskeleton-dependent manner, depending on intact actin and myosin ATP-hydrolysis, and is largely bound to the actin cytoskeleton. The ZBP1 protein contains four KH RNA binding domains and a classic RBD RNA binding domain. It also contains a putative nuclear import and export sequence, suggesting a nuclear phase in this protein's function. RESULTS:Using high-speed imaging, we show here the targeting of this RNA binding protein to beta-actin pre-mRNA transcripts in the nuclei of living cells and measure the residence time of the RNA-protein complex before it leaves the transcription site. Then, the RNA-protein particle is exported to the cytoplasm, where it localizes at velocities of 0.6 microm/s by using actin filaments and/or microtubules. This RNA-ZBP1 complex is required for cytoplasmic localization in fibroblasts; mislocalizing the protein also mislocalizes the RNA, and expressing the protein in a ZBP1-deficient cell line induces beta-actin mRNA localization. CONCLUSIONS:This work demonstrates that the RNA-protein association, essential for cytoplasmic localization, begins as soon as the RNA is transcribed. The ZBP1 then forms a ribonucleoprotein particle and moves in a myosin-dependent fashion by using the cytoskeleton for directional transport.

| S-EPMC4765734 | biostudies-literature

Specific interaction of KIF11 with ZBP1 regulates the transport of ?-actin mRNA and cell motility.

Project description:ZBP1-modulated localization of ?-actin mRNA enables a cell to establish polarity and structural asymmetry. Although the mechanism of ?-actin mRNA localization has been well established, the underlying mechanism of how a specific molecular motor contributes to the transport of the ZBP1 (also known as IGF2BP1) complex in non-neuronal cells remains elusive. In this study, we report the isolation and identification of KIF11, a microtubule motor, which physically interacts with ZBP1 and is a component of ?-actin messenger ribonucleoprotein particles (mRNPs). We show that KIF11 colocalizes with the ?-actin mRNA, and the ability of KIF11 to transport ?-actin mRNA is dependent on ZBP1. We characterize the corresponding regions of ZBP1 and KIF11 that mediate the interaction of the two proteins in vitro and in vivo. Disruption of the in vivo interaction of KIF11 with ZBP1 delocalizes ?-actin mRNA and affects cell migration. Our study reveals a molecular mechanism by which a particular microtubule motor mediates the transport of an mRNP through direct interaction with an mRNA-binding protein.

| S-EPMC4342582 | biostudies-literature

Different motif requirements for the localization zipcode element of β-actin mRNA binding by HuD and ZBP1.

Project description:Interactions of RNA-binding proteins (RBPs) with their target transcripts are essential for regulating gene expression at the posttranscriptional level including mRNA export/localization, stability, and translation. ZBP1 and HuD are RBPs that play pivotal roles in mRNA transport and local translational control in neuronal processes. While HuD possesses three RNA recognition motifs (RRMs), ZBP1 contains two RRMs and four K homology (KH) domains that either increase target specificity or provide a multi-target binding capability. Here we used isolated cis-element sequences of the target mRNA to examine directly protein-RNA interactions in cell-free systems. We found that both ZBP1 and HuD bind the zipcode element in rat β-actin mRNA's 3' UTR. Differences between HuD and ZBP1 were observed in their binding preference to the element. HuD showed a binding preference for U-rich sequence. In contrast, ZBP1 binding to the zipcode RNA depended more on the structural level, as it required the proper spatial organization of a stem-loop that is mainly determined by the U-rich element juxtaposed to the 3' end of a 5'-ACACCC-3' motif. On the basis of this work, we propose that ZBP1 and HuD bind to overlapping sites in the β-actin zipcode, but they recognize different features of this target sequence.

| S-EPMC4551932 | biostudies-literature

ZBP1 regulates mRNA stability during cellular stress.

Project description:An essential constituent of the integrated stress response (ISR) is a reversible translational suppression. This mRNA silencing occurs in distinct cytoplasmic foci called stress granules (SGs), which transiently associate with processing bodies (PBs), typically serving as mRNA decay centers. How mRNAs are protected from degradation in these structures remains elusive. We identify that Zipcode-binding protein 1 (ZBP1) regulates the cytoplasmic fate of specific mRNAs in nonstressed cells and is a key regulator of mRNA turnover during the ISR. ZBP1 association with target mRNAs in SGs was not essential for mRNA targeting to SGs. However, ZBP1 knockdown induced a selective destabilization of target mRNAs during the ISR, whereas forced expression increased mRNA stability. Our results indicate that although targeting of mRNAs to SGs is nonspecific, the stabilization of mRNAs during cellular stress requires specific protein-mRNA interactions. These retain mRNAs in SGs and prevent premature decay in PBs. Hence, mRNA-binding proteins are essential for translational adaptation during cellular stress by modulating mRNA turnover.

| S-EPMC2064588 | biostudies-literature

ZBP1 condensate formation synergizes Z-NAs recognition and signal transduction.

Project description:Z-DNA binding protein 1 (ZBP1) is a crucial player in the intracellular recognition of Z-form nucleic acids (Z-NAs) through its Zαβ domain, initiating downstream interactions with RIPK1 and RIPK3 via RHIM domains. This engagement leads to the assembly of PANoptosomes, ultimately inducing programmed cell death to curb pathogen dissemination. How Zαβ and RHIM domain cooperate to trigger Z-NAs recognition and signal transduction remains unclear. Here, we show that ZBP1 condensate formation facilitates Z-NAs binding and antiviral signal transduction. The ZBP1 Zαβ dimerizes in a concentration-dependent manner, forming characteristic condensates in solutions evidenced by DLS and SAXS methods. ZBP1 exhibits a binding preference for 10-bp length CG (10CG) DNA and Z-RNA ligand, which in turn enhanced Zαβ dimerization, expediting the formation of droplet condensates in vitro and amyloid-like puncta in cells. Subsequent investigations reveal that Zαβ could form condensates with liquid-liquid phase separation property upon HSV and IAV infections, while full-length ZBP1 forms amyloid-like puncta with or without infections. Furthermore, ZBP1 RHIM domains show typical amyloidal fibril characterizations and cross-polymerize with RIPK1 depending on the core motif of 206IQIG209, while mutated ZBP1 could impede necroptosis and antiviral immunity in HT-29 cells. Thus, ZBP1 condensate formation facilitates the recognition of viral Z-NAs and activation of downstream signal transduction via synergic action of different domains, revealing its elaborated mechanism in innate immunity.

| S-EPMC11233663 | biostudies-literature

Single-molecule analysis of endogenous ?-actin mRNA trafficking reveals a mechanism for compartmentalized mRNA localization in axons.

Project description:During embryonic nervous system assembly, mRNA localization is precisely regulated in growing axons, affording subcellular autonomy by allowing controlled protein expression in space and time. Different sets of mRNAs exhibit different localization patterns across the axon. However, little is known about how mRNAs move in axons or how these patterns are generated. Here, we couple molecular beacon technology with highly inclined and laminated optical sheet microscopy to image single molecules of identified endogenous mRNA in growing axons. By combining quantitative single-molecule imaging with biophysical motion models, we show that ?-actin mRNA travels mainly as single copies and exhibits different motion-type frequencies in different axonal subcompartments. We find that ?-actin mRNA density is fourfold enriched in the growth cone central domain compared with the axon shaft and that a modicum of directed transport is vital for delivery of mRNA to the axon tip. Through mathematical modeling we further demonstrate that directional differences in motor-driven mRNA transport speeds are sufficient to generate ?-actin mRNA enrichment at the growth cone. Our results provide insight into how mRNAs are trafficked in axons and a mechanism for generating different mRNA densities across axonal subcompartments.

| S-EPMC6187124 | biostudies-literature

Limited availability of ZBP1 restricts axonal mRNA localization and nerve regeneration capacity.

Project description:Subcellular localization of mRNAs is regulated by RNA-protein interactions. Here, we show that introduction of a reporter mRNA with the 3'UTR of ?-actin mRNA competes with endogenous mRNAs for binding to ZBP1 in adult sensory neurons. ZBP1 is needed for axonal localization of ?-actin mRNA, and introducing GFP with the 3'UTR of ?-actin mRNA depletes axons of endogenous ?-actin and GAP-43 mRNAs and attenuates both in vitro and in vivo regrowth of severed axons. Consistent with limited levels of ZBP1 protein in adult neurons, mice heterozygous for the ZBP1 gene are haploinsufficient for axonal transport of ?-actin and GAP-43 mRNAs and for regeneration of peripheral nerve. Exogenous ZBP1 can rescue the RNA transport deficits, but the axonal growth deficit is only rescued if the transported mRNAs are locally translated. These data support a direct role for ZBP1 in transport and translation of mRNA cargos in axonal regeneration in vitro and in vivo.

| S-EPMC3243598 | biostudies-literature

OmicsDI is part of the ELIXIR infrastructure

OmicsDI is an Elixir interoperability service. Learn more ›

Tweets

OmicsDI Databases

PRIDE
PeptideAtlas
MassIVE
JPOST Repository
Physiome Model Repository

EGA
EVA
ENA
LINCS
PAXDB
Cell Collective

MetaboLights
Metabolomics Workbench
MetabolomeExpress
GNPS
BioModels
FAIRDOMHub

ArrayExpress
dbGaP
ExpressionAtlas
GEO
NODE

Information

Databases
Help
API
Contact us
Code on GitHub
Terms of use
Submit Data