yRACK1/Asc1 proxiOMICs—Towards Illuminating Ships Passing in the Night

Diverse signals and stress factors regulate the activity and homeostasis of ribosomes in all cells. The Saccharomyces cerevisiae protein Asc1/yRACK1 occupies an exposed site at the head region of the 40S ribosomal subunit (hr40S) and represents a central hub for signaling pathways. Asc1 strongly affects protein phosphorylation and is involved in quality control pathways induced by translation elongation arrest. Therefore, it is important to understand the dynamics of protein formations in the Asc1 microenvironment at the hr40S. We made use of the in vivo protein-proximity labeling technique Biotin IDentification (BioID). Unbiased proxiOMICs from two adjacent perspectives identified nucleocytoplasmic shuttling mRNA-binding proteins, the deubiquitinase complex Ubp3-Bre5, as well as the ubiquitin E3 ligase Hel2 as neighbors of Asc1. We observed Asc1-dependency of hr40S localization of mRNA-binding proteins and the Ubp3 co-factor Bre5. Hel2 and Ubp3-Bre5 are described to balance the mono-ubiquitination of Rps3 (uS3) during ribosome quality control. Here, we show that the absence of Asc1 resulted in massive exposure and accessibility of the C-terminal tail of its ribosomal neighbor Rps3 (uS3). Asc1 and some of its direct neighbors together might form a ribosomal decision tree that is tightly connected to close-by signaling modules.

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Direct Link between RACK1 Function and Localization at the Ribosome In Vivo

Molecular and Cellular Biology ◽

10.1128/mcb.01718-08 ◽

2008 ◽

Vol 29 (6) ◽

pp. 1626-1634 ◽

Cited By ~ 107

Author(s):

Scott M. Coyle ◽

Wendy V. Gilbert ◽

Jennifer A. Doudna

Keyword(s):

Signaling Pathways ◽

Ribosomal Subunit ◽

Translation Elongation ◽

Ribosome Binding ◽

Charged Amino Acids ◽

40S Ribosomal Subunit ◽

Increased Sensitivity ◽

Mrna Binding ◽

Ribosome Association

ABSTRACT The receptor for activated C-kinase (RACK1), a conserved protein implicated in numerous signaling pathways, is a stoichiometric component of eukaryotic ribosomes located on the head of the 40S ribosomal subunit. To test the hypothesis that ribosome association is central to the function of RACK1 in vivo, we determined the 2.1-Å crystal structure of RACK1 from Saccharomyces cerevisiae (Asc1p) and used it to design eight mutant versions of RACK1 to assess roles in ribosome binding and in vivo function. Conserved charged amino acids on one side of the β-propeller structure were found to confer most of the 40S subunit binding affinity, whereas an adjacent conserved and structured loop had little effect on RACK1-ribosome association. Yeast mutations that confer moderate to strong defects in ribosome binding mimic some phenotypes of a RACK1 deletion strain, including increased sensitivity to drugs affecting cell wall biosynthesis and translation elongation. Furthermore, disruption of RACK1's position at the 40S ribosomal subunit results in the failure of the mRNA binding protein Scp160 to associate with actively translating ribosomes. These results provide the first direct evidence that RACK1 functions from the ribosome, implying a physical link between the eukaryotic ribosome and cell signaling pathways in vivo.

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In Vivo Cross-Linking Followed by PolyA Enrichment to Identify Yeast mRNA Binding Proteins

Methods in Molecular Biology - RNA Remodeling Proteins ◽

10.1007/978-1-4939-2214-7_3 ◽

2014 ◽

pp. 35-47 ◽

Cited By ~ 1

Author(s):

Sarah F. Mitchell ◽

Roy Parker

Keyword(s):

Binding Proteins ◽

Cross Linking ◽

Mrna Binding Proteins ◽

Mrna Binding

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In Vivo Cross-Linking Followed by polyA Enrichment to Identify Yeast mRNA Binding Proteins

Methods in Molecular Biology - RNA Remodeling Proteins ◽

10.1007/978-1-0716-0935-4_15 ◽

2020 ◽

pp. 235-249

Author(s):

Sarah F. Mitchell

Keyword(s):

Binding Proteins ◽

Cross Linking ◽

Mrna Binding Proteins ◽

Mrna Binding

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Modification of mRNA-associated proteins during changes in growth conditions

Biochemistry and Cell Biology ◽

10.1139/o92-082 ◽

1992 ◽

Vol 70 (7) ◽

pp. 528-534 ◽

Cited By ~ 2

Author(s):

Lloyd C. Berger ◽

Bruce H. Sells

Keyword(s):

Binding Proteins ◽

Growth Conditions ◽

Serum Stimulation ◽

Mrna Binding Proteins ◽

Polysomal Mrna ◽

Associated Proteins ◽

Growth Transitions ◽

Mrna Binding ◽

Stimulation Of

Following serum stimulation of quiescent 3T6 cells, an elevated in vivo rate of translation was observed. These studies were designed to identify the proteins associated with polysomal mRNA under different growth conditions in an attempt to establish a relationship between translational rate and the mRNA-associated proteins. Ultraviolet cross-linking of proteins to mRNA was employed to ensure that only genuine mRNA-associated proteins were investigated. Our results revealed little change in the population of mRNA-binding proteins, although minor variations in the synthesis of several proteins, most notably a 32 kilodalton species, were observed during growth transitions. These investigations demonstrate further that most of the mRNA-binding proteins were phosphorylated with the degree of phosphorylation of several proteins influenced by growth conditions.Key words: mRNA-associated proteins, phosphorylation, translation.

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Faculty Opinions recommendation of Insights into RNA biology from an atlas of mammalian mRNA-binding proteins.

Faculty Opinions – Post-Publication Peer Review of the Biomedical Literature ◽

10.3410/f.717949150.793524523 ◽

2016 ◽

Author(s):

Sean Ryder

Keyword(s):

Binding Proteins ◽

Mrna Binding Proteins ◽

Rna Biology ◽

Mrna Binding

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The endoplasmic reticulum-associated mRNA-binding proteins ERBP1 and ERBP2 interact in bloodstream-form Trypanosoma brucei

10.1101/718031 ◽

2019 ◽

Author(s):

Kathrin Bajak ◽

Kevin Leiss ◽

Christine Clayton ◽

Esteban Erben

Keyword(s):

Endoplasmic Reticulum ◽

Ribosomal Protein ◽

Trypanosoma Brucei ◽

Binding Proteins ◽

Rna Binding ◽

Rna Binding Proteins ◽

Bloodstream Form ◽

Control Of Gene Expression ◽

Mrna Binding Proteins ◽

Mrna Binding

AbstractKinetoplastids rely heavily on post-transcriptional mechanisms for control of gene expression, and on RNA-binding proteins that regulate mRNA splicing, translation and decay. Trypanosoma brucei ERBP1 (Tb927.10.14150) and ERBP2 (Tb927.9.9550) were previously identified as mRNA binding proteins that lack canonical RNA-binding domains. We here show that ERBP1 is associated with the endoplasmic reticulum, like ERBP2, and that the two proteins interact in vivo. Loss of ERBP1 from bloodstream-form T. brucei initially resulted in a growth defect but proliferation was restored after more prolonged cultivation. Results from a pull-down of tagged ERBP1 suggest that it preferentially binds to ribosomal protein mRNAs. The ERBP1 sequence resembles that of Saccharomyces cerevisiae Bfr1, which also localises to the endoplasmic reticulum and binds to ribosomal protein mRNAs. However, unlike Bfr1, ERBP1 does not bind to mRNAs encoding secreted proteins, and it is also not recruited to stress granules after starvation.

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Humoral Autoimmune Responses to Insulin-Like Growth Factor II mRNA-Binding Proteins IMP1 and p62/IMP2 in Ovarian Cancer

Journal of Immunology Research ◽

10.1155/2014/326593 ◽

2014 ◽

Vol 2014 ◽

pp. 1-7 ◽

Cited By ~ 5

Author(s):

Xinxin Liu ◽

Hua Ye ◽

Liuxia Li ◽

Wenjie Li ◽

Yi Zhang ◽

...

Keyword(s):

Ovarian Cancer ◽

Growth Factor ◽

Binding Proteins ◽

Immunofluorescence Assay ◽

Enzyme Linked Immunosorbent Assay ◽

Insulin Like Growth Factor ◽

Mrna Binding Proteins ◽

Normal Individuals ◽

Factor Ii ◽

Mrna Binding

Ovarian cancer is one of the leading causes of cancer-related deaths among women. There is an urgent need of better approaches for the identification of appropriate biomarkers in the early detection of ovarian cancer. The aim of this study was to elucidate the significance of autoantibodies against insulin-like growth factor II mRNA-binding proteins (IMPs) in patients with ovarian cancer. In this study, autoantibody responses to two members (IMP1 and p62/IMP2) of IMPs were evaluated by enzyme-linked immunosorbent assay (ELISA), western blotting, and indirect immunofluorescence assay in sera from patients with ovarian cancer and normal human individuals. The results have demonstrated that both IMP1 and p62/IMP2 can induce relatively higher frequency of autoantibody responses in patients with ovarian cancer (26.5% and 29.4%) compared to normal individuals(P<0.01). Our preliminary data suggest that IMP1 and p62/IMP2 can stimulate autoimmune responses in ovarian cancer, and anti-IMP1 and anti-p62/IMP2 autoantibodies could be used as potential biomarkers in immunodiagnosis of ovarian cancer.

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