Dataset Information

RNA co-immunoprecipitation coupled to RNA sequencing (RIP-seq) to identify somatic target mRNAs of the TRIM-NHL protein LIN41

ABSTRACT: We perform RIP-seq experiments with two C. elegans worm lines: i) lin-41(n2914); him-5(e1490) with transgenic expression of a rescuing flag::gfp::lin-41 transgene (Aeschimann et al., Mol Cell, 2017) and ii) him-5(e1490) with transgenic expression of flag::gfp::sart-3 (Rüegger et al., NAR, 2015) as a control. The him-5(e1490) genetic background results in an increased frequency of males in the population. We used an anti-FLAG antibody for the IP and semi-synchronous populations of animals in the L3 and L4 larval stages as samples. The purpose of the experiment was to identify LIN41 mRNA targets in the soma. From the three independent replicates, we determined a set of LIN41-bound mRNAs using edgeR and FDR < 0.05 as a cutoff. This set contained only seven mRNAs, the previously known targets lin-29, mab-10, mab-3 and dmd-3 (Aeschimann et al., Mol Cell, 2017), as well as lin-41 and the unannotated genes F18C5.10 and C31H5.5. We conclude that LIN41 likely only binds to a few somatic mRNA targets. Intersecting this experiment with differential gene expression experiments upon dys-regulation of LIN41 (Aeschimann et al., Mol Cell, 2017) and phenotypic analysis of mutant strains, we further conclude that during larval development, LIN41 likely regulates only a four direct targets, namely lin-29, mab-10, mab-3 and dmd-3. Additionally, the lin-41 mRNA could be directly targeted by LIN41 as well, but the detection of lin-41 mRNA in the IPs may result from the immunoprecipitation of nascent FLAG::GFP::LIN41 protein, still bound to the translating ribosome on its own mRNA.

ORGANISM(S): Caenorhabditis elegans

PROVIDER: GSE120405 | GEO | 2018/11/15

REPOSITORIES: GEO

ACCESS DATA

Dataset's files

Source:

			Action	DRS
		Other

Items per page:

1 - 1 of 1

Similar Datasets

Project description:CD47 is a transmembrane glycoprotein that is ubiquitously expressed in different organs and tissues (Barclay and Van den Berg 2014; Liu, et al. 2017). In the human immune system, CD47 interacts with some integrins, two counter-receptor signal regulator protein (SIRP) family members, and the secreted thrombospondin-1 (TSP1) (Barclay and Van den Berg 2014; Gao, et al. 2016; Kaur, et al. 2013; Oldenborg, et al. 2000). CD47 has two established roles in the immune system. The CD47-SIRPα interaction was identified as a critical innate immune checkpoint, which delivers an antiphagocytic signal to macrophages and inhibits neutrophil cytotoxicity (Martínez- Sanz, et al. 2021). Its interaction with inhibitory SIRPα is a physiological anti-phagocytic “don’t eat me” signal on circulating red blood cells that is co-opted by cancer cells (Matlung, et al. 2017). Many malignant cells overexpress CD47 (Betancur, et al. 2017; Chao, et al. 2011; Jaiswal, et al. 2009; Majeti, et al. 2009; Oronsky, et al. 2020; Petrova, et al. 2017). CD47/SIRPα-targeted therapeutics have been developed to overcome this immune checkpoint for cancer treatment (Kaur, et al. 2020; Matlung, et al. 2017). Secondly, engagement of CD47 on T cells by TSP1 regulates their differentiation and survival (Grimbert, et al. 2006; Lamy, et al. 2007) and inhibits T cell receptor signaling and antigen presentation by dendritic cells (DCs) (Kaur, et al. 2014; Li, et al. 2002; Liu, et al. 2015; Miller, et al. 2013; Soto-Pantoja, et al. 2014; Weng, et al. 2014). TSP1/CD47 signaling has similar inhibitory functions to limit NK cell activation (Kim, et al. 2008; Nath, et al. 2018; Nath, et al. 2019; Schwartz, et al. 2019) and IL1β production by macrophages (Stein, et al. 2016). CD47 is therefore a checkpoint that regulates both innate and adaptive immunity. The recent understanding of CD47 antagonism associated with increased antigen presentation by DCs (Liu, et al. 2016) and natural killer cell cytotoxicity (Nath, et al. 2019) contributes to the heightened interest in CD47 as a therapeutic target (Kaur, et al. 2020).

Project description:Many tissue-specific stem cells maintain the ability to produce multiple cell types during long periods of non-division, or quiescence. FOXO transcription factors promote quiescence and stem cell maintenance, but the mechanisms by which FOXO proteins promote multipotency during quiescence are still emerging. The single FOXO ortholog in C. elegans, daf-16, promotes entry into a quiescent and stress-resistant larval stage called dauer in response to adverse environmental cues. During dauer, stem and progenitor cells maintain or re-establish multipotency to allow normal development to resume after dauer. We find that during dauer, daf-16/FOXO prevents epidermal stem cells (seam cells) from prematurely adopting differentiated, adult characteristics. In particular, dauer larvae that lack daf-16 misexpress collagens that are normally adult-enriched. Using col-19p::gfp as an adult cell fate marker, we find that all major daf-16 isoforms contribute to opposing col-19p::gfp expression during dauer. By contrast, daf-16(0) larvae that undergo non-dauer development do not misexpress col-19p::gfp. Adult cell fate and the timing of col-19p::gfp expression are regulated by the heterochronic gene network, including lin-41 and lin-29. lin-41 encodes an RNA-binding protein orthologous to LIN41/TRIM71 in mammals, and lin-29 encodes a conserved zinc finger transcription factor. In non-dauer development lin-41 opposes adult cell fate by inhibiting the translation of lin-29, which directly activates col-19 transcription and promotes adult cell fate. We find that during dauer, lin-41 blocks col-19p::gfp expression, but surprisingly, lin-29 is not required in this context. Additionally, daf-16 promotes the expression of lin-41 in dauer larvae. The col-19p::gfp misexpression phenotype observed in dauer larvae with reduced daf-16 requires the downregulation of lin-41, but does not require lin-29. Taken together, this work demonstrates a novel role for daf-16/FOXO as a heterochronic gene that promotes expression of lin-41/TRIM71 to contribute to multipotent cell fate in a quiescent stem cell model.

Dataset Information

RNA co-immunoprecipitation coupled to RNA sequencing (RIP-seq) to identify somatic target mRNAs of the TRIM-NHL protein LIN41

Dataset's files

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets