A functional screen of RNA binding proteins identifies genes that promote or limit the accumulation of CD138+ plasma cells
- The Babraham Institute, United Kingdom
- Queen Mary University of London, United Kingdom
Abstract
To identify roles of RNA binding proteins (RBPs) in the differentiation or survival of antibody secreting plasma cells we performed a CRISPR/Cas9 knockout screen of 1213 mouse RBPs for their ability to affect proliferation and/or survival, and the abundance of differentiated CD138+ cells in vitro. We validated the binding partners CSDE1 and STRAP as well as the m6A binding protein YTHDF2 as promoting the accumulation of CD138+ cells in vitro. We validated the EIF3 subunits EIF3K and EIF3L and components of the CCR4-NOT complex as inhibitors of CD138+ cell accumulation in vitro. In chimeric mouse models YTHDF2-deficient plasma cells failed to accumulate.
Data availability
The sgRNA library is available upon request and from Addgene (#169082). The CRISPR/Cas9 knockout screen data and m6A-eCLIP data that support the findings of this study have been deposited in GEO with the GSE179919 accession code, and the RNA-seq data has been deposited in GEO with the GSE179281 accession code.
-
The RNA m6A binding protein YTHDF2 promotes the B cell to plasma cell transitionNCBI Gene Expression Omnibus, GSE179919.
-
Transcriptomic analysis of in vitro induced germinal centre-like B cells and plasmablast differentiationNCBI Gene Expression Omnibus, GSE179281.
Article and author information
Author details
Funding
Biotechnology and Biological Sciences Research Council (BBS/E/B/000C0427)
- Martin Turner
Biotechnology and Biological Sciences Research Council (BBS/E/B/000C0428)
- Martin Turner
Wellcome Trust (200823/Z/16/Z)
- Martin Turner
Biotechnology and Biological Sciences Research Council (BB/L016745/1)
- David J Turner
European Molecular Biology Organisation (ALTF 880-2018)
- Fiamma Salerno
Cancer Research UK (C29967/A26787)
- Kamil R Kranc
The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.
Reviewing Editor
- Tomohiro Kurosaki, Osaka University, Japan
Ethics
Animal experimentation: All mouse experimentation was approved by the Babraham Institute Animal Welfare and Ethical Review Body and was licensed by the United Kingdom Home Office under PPL P4D4AF812.
Version history
- Received: July 19, 2021
- Preprint posted: July 21, 2021 (view preprint)
- Accepted: April 21, 2022
- Accepted Manuscript published: April 22, 2022 (version 1)
- Version of Record published: May 13, 2022 (version 2)
Copyright
© 2022, Turner et al.
This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.
Metrics
-
- 2,973
- views
-
- 361
- downloads
-
- 5
- citations
Views, downloads and citations are aggregated across all versions of this paper published by eLife.
Download links
A two-part list of links to download the article, or parts of the article, in various formats.
Downloads (link to download the article as PDF)
Open citations (links to open the citations from this article in various online reference manager services)
Cite this article (links to download the citations from this article in formats compatible with various reference manager tools)
A functional screen of RNA binding proteins identifies genes that promote or limit the accumulation of CD138+ plasma cells
eLife 11:e72313.
https://doi.org/10.7554/eLife.72313
Further reading
-
- Developmental Biology
Precise developmental timing control is essential for organism formation and function, but its mechanisms are unclear. In C. elegans, the microRNA lin-4 critically regulates developmental timing by post-transcriptionally downregulating the larval-stage-fate controller LIN-14. However, the mechanisms triggering the activation of lin-4 expression toward the end of the first larval stage remain unknown. We demonstrate that the transmembrane transcription factor MYRF-1 is necessary for lin-4 activation. MYRF-1 is initially localized on the cell membrane, and its increased cleavage and nuclear accumulation coincide with lin-4 expression timing. MYRF-1 regulates lin-4 expression cell-autonomously and hyperactive MYRF-1 can prematurely drive lin-4 expression in embryos and young first-stage larvae. The tandem lin-4 promoter DNA recruits MYRF-1GFP to form visible loci in the nucleus, suggesting that MYRF-1 directly binds to the lin-4 promoter. Our findings identify a crucial link in understanding developmental timing regulation and establish MYRF-1 as a key regulator of lin-4 expression.
-
- Developmental Biology
- Structural Biology and Molecular Biophysics
The receptor tyrosine kinase ROR2 mediates noncanonical WNT5A signaling to orchestrate tissue morphogenetic processes, and dysfunction of the pathway causes Robinow syndrome, brachydactyly B, and metastatic diseases. The domain(s) and mechanisms required for ROR2 function, however, remain unclear. We solved the crystal structure of the extracellular cysteine-rich (CRD) and Kringle (Kr) domains of ROR2 and found that, unlike other CRDs, the ROR2 CRD lacks the signature hydrophobic pocket that binds lipids/lipid-modified proteins, such as WNTs, suggesting a novel mechanism of ligand reception. Functionally, we showed that the ROR2 CRD, but not other domains, is required and minimally sufficient to promote WNT5A signaling, and Robinow mutations in the CRD and the adjacent Kr impair ROR2 secretion and function. Moreover, using function-activating and -perturbing antibodies against the Frizzled (FZ) family of WNT receptors, we demonstrate the involvement of FZ in WNT5A-ROR signaling. Thus, ROR2 acts via its CRD to potentiate the function of a receptor super-complex that includes FZ to transduce WNT5A signals.
