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Post-transcriptional gene regulation in development: the function of LIN-41/TRIM71

English title Post-transcriptional gene regulation in development: the function of LIN-41/TRIM71
Applicant Grosshans Helge
Number 163447
Funding scheme Project funding (Div. I-III)
Research institution Friedrich Miescher Institute for Biomedical Research
Institution of higher education Institute Friedrich Miescher - FMI
Main discipline Molecular Biology
Start/End 01.06.2016 - 31.10.2019
Approved amount 756'000.00
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Keywords (9)

translational control; developmental timing; LIN-41; TRIM71; TRIM-NHL; C. elegans; posttranscriptional gene regulation; stem cell; RNA turnover

Lay Summary (German)

Lead
Die Aktivität von Genen muss räumlich und zeitlich genau reguliert werden, damit sich Organismen entwickeln können. Daher ist nicht nur das Ablesen der Gene strikt kontrolliert, sondern auch die weitere Verwendung der entstandenen Abschriften. In verschiedenen Stammzellen verhindert LIN-41/TRIM71, dass Abschriften für die Produktion von Protein verwendet werden. Die zugrundeliegenden Mechanismen sind nur ansatzweise verstanden.
Lay summary

Wir haben kürzlich im Fadenwurm Caenorhabditis elegans gezeigt, dass LIN-41/TRIM71 zentral für die Regulation verschiedener Entwicklungsprozesse ist. Dieses Protein existiert auch in Säugetieren einschliesslich des Menschen und reguliert dort wie im Wurm die besonderen Eigenschaften von Stammzellen. Unser Ziel ist, im Detail zu verstehen, wie LIN-41/TRIM71 funktioniert. Wir erforschen insbesondere seine molekularen Funktionsweise sowie seine spezifischen Zielgene. Zu diesem Zweck werden wir genetische, biochemische, genom-basierte und zellbiologische Methoden in C. elegans anwenden.

 

Wissenschaftlicher und gesellschaftlicher Kontext des Forschungsprojekts

Unsere Arbeit zielt darauf ab, neue Einsichten in die Komplexität und Dynamik der Genkontrolle sowie in die Regulation von Stammzellen eröffnen. Damit vertiefen wir das Verständnis der molekularen Mechanismen, die tierische Entwicklungsprozesse befördern. Dieses Wissen wird beispielsweise in der regenerativen Medizin für die Entwicklung neuartiger Therapieansätze benötigt.

Direct link to Lay Summary Last update: 04.12.2015

Responsible applicant and co-applicants

Employees

Publications

Publication
let-7 coordinates the transition to adulthood through a single primary and four secondary targets
Aeschimann Florian, Neagu Anca, Rausch Magdalene, Großhans Helge (2019), let-7 coordinates the transition to adulthood through a single primary and four secondary targets, in Life Science Alliance, 2(2), e201900335-e201900335.
Evolutionary plasticity of the NHL domain underlies distinct solutions to RNA recognition
Kumari Pooja, Aeschimann Florian, Gaidatzis Dimos, Keusch Jeremy J., Ghosh Pritha, Neagu Anca, Pachulska-Wieczorek Katarzyna, Bujnicki Janusz M., Gut Heinz, Großhans Helge, Ciosk Rafal (2018), Evolutionary plasticity of the NHL domain underlies distinct solutions to RNA recognition, in Nature Communications, 9(1), 1549-1549.
LIN41 Post-transcriptionally Silences mRNAs by Two Distinct and Position-Dependent Mechanisms.
Aeschimann Florian, Kumari Pooja, Bartake Hrishikesh, Gaidatzis Dimos, Xu Lan, Ciosk Rafal, Großhans Helge (2017), LIN41 Post-transcriptionally Silences mRNAs by Two Distinct and Position-Dependent Mechanisms., in Molecular cell, 65(3), 476-489.

Datasets

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

Author Aeschimann, Florian
Publication date 15.11.2018
Persistent Identifier (PID) GSE120405
Repository GEO
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.

LIN-41 RIP-Seq

Author Kumari, Pooja
Publication date 25.04.2018
Persistent Identifier (PID) GSE106814
Repository GEO
Abstract
FLAG-GFP tagged LIN-41 was immunoprecipitated using FLAG dynabeads (Sigma) from lysates of young adult C.elegans. After washing steps, bound fraction (IP) was eluted from beads and associated RNAs were isolated by using the Pico pure RNA isloation kit (Invitrogen)

Post-transcriptional regulation by the let-7 microRNA and the TRIM-NHL protein LIN41 [RNA-seq]

Author Aeschimann, Florian
Publication date 17.07.2017
Persistent Identifier (PID) GSE80157
Repository GEO
Abstract
We perform RNA sequencing and ribosome profiling time course experiments to examine the effect of fully dysregulating all let-7 targets (in let-7(n2853) animals), partially dysregulating only LIN41 (in lin-41(xe11) animals) or fully dysregulating all let-7 targets while partially dysregulating LIN41 in lin-41(xe11); let-7(n2853) double mutant animals. We conclude that effects on gene expression in let-7 mutant animals are largely and quantitatively explained by dysregulation of LIN41 as its primary target. Furthermore, we identify direct LIN41 target genes regulated on the level of translation or mRNA abundance.

Post-transcriptional regulation by the let-7 microRNA and the TRIM-NHL protein LIN41 [Ribosome footprinting]

Author Aeschimann, Florian
Publication date 17.07.2017
Persistent Identifier (PID) GSE80133
Repository GEO
Abstract
We perform RNA sequencing and ribosome profiling time course experiments to examine the effect of fully dysregulating all let-7 targets (in let-7(n2853) animals), partially dysregulating only LIN41 (in lin-41(xe11) animals) or fully dysregulating all let-7 targets while partially dysregulating LIN41 in lin-41(xe11); let-7(n2853) double mutant animals. We conclude that effects on gene expression in let-7 mutant animals are largely and quantitatively explained by dysregulation of LIN41 as its primary target. Furthermore, we identify direct LIN41 target genes regulated on the level of translation or mRNA abundance.

Collaboration

Group / person Country
Types of collaboration
Dr. Rafal Ciosk, FMI Switzerland (Europe)
- in-depth/constructive exchanges on approaches, methods or results
- Publication
Dr. Oliver Hobert United States of America (North America)
- in-depth/constructive exchanges on approaches, methods or results
- Publication

Scientific events

Active participation

Title Type of contribution Title of article or contribution Date Place Persons involved
22nd International C. elegans Conference Talk given at a conference Two parallel arms of the heterochronic pathway direct coordinated juvenile-to-adult transition through distinct LIN-29 isoforms 20.06.2019 Los Angeles, CA, United States of America Azzi Chiarra;
Keystone Symposium “Small Regulatory RNAs” Talk given at a conference Promoting Adulthood Through a miRNA With a Single Target 14.04.2019 Daejeon, Korean Republic (South Korea) Grosshans Helge;
Basel Worm Meeting 2019 Talk given at a conference C.elegans progenitor cell fate control through the EGR-type transcription factor LIN-29 28.03.2019 Basel, Switzerland Azzi Chiarra;
Swiss RNA Workshop Poster Molecular and developmental functions of mRNA silencing by LIN41 25.01.2019 Bern, Switzerland Liu Jun;
EMBO | EMBL Symposium: The Complex Life of mRNA Poster Molecular and developmental functions of mRNA silencing by LIN41 03.10.2018 Heidelberg, Germany Liu Jun;
European Worm Meeting Poster C.elegans progenitor cell fate control through the EGR-type transcription factor LIN-29 13.06.2018 Barcelona, Spain Azzi Chiarra;
EMBO Workshop: C. elegans Development, Cell Biology, and Gene Expression Poster The microRNA let-7 controls three distinct developmental events exclusively through the RNA-binding protein LIN41 and its four targets 13.06.2018 Barcelona, Spain Aeschimann Florian;
Cancer, Stem Cells, and Developmental Biology Graduate Programme Course in Developmental Genetics Individual talk Timing development with biological oscillators and 
post-transcriptional timers 18.04.2018 Utrecht University/Hubrecht Institute, Netherlands Grosshans Helge;
Cologne Spring Meeting 2018 Talk given at a conference Post-Transcriptional Regulation of Cell Fates by 
let-7 & LIN41/TRIM71 07.03.2018 Köln, Germany Grosshans Helge;
Seminar Individual talk Cell fate control through posttranscriptional regulation 
and clocks 29.09.2017 Laval University, Montreal, Canada Grosshans Helge;
The Non-Coding Genome (EMBO Conference Series) Poster The RNA-binding protein LIN41 binds to 3’ and 5’ untranslated regions to silence mRNAs through distinct mechanism 13.09.2017 Heidelberg, Germany Aeschimann Florian;
EMBO at Basel Life Talk given at a conference Regulation of progenitor cell fates through RNA 10.09.2017 Basel, Switzerland Grosshans Helge;
EMBO at Basel Life Poster The RNA-binding protein LIN41 silences mRNAs through distinct mechanisms depending on its binding location 10.09.2017 Basel, Switzerland Aeschimann Florian;
ISFMS 2017: Non-Coding RNAs and Epigenetics in Cancer Talk given at a conference Noncoding RNA Function and Regulation in Animal Development 21.06.2017 Basel, Switzerland Grosshans Helge;
Seminar Individual talk Cell fate control through biological clocks 
and post-transcriptional timekeeping 18.04.2017 IMP, Vienna, Austria Grosshans Helge;
3rd Curie International Course on Post-Transcriptional Gene Regulation Individual talk Post-Transcriptional Mechanisms that Control Cell Fates and Developmental Timing 30.03.2017 Paris, France Grosshans Helge;
3rd Curie International Course on Post-Transcriptional Gene Regulation Poster Mechanistic and developmental functions of LIN41/TRIM71 as a post-transcriptional silencer in C. elegans 27.03.2017 Paris, France Liu Jun;
Cambridge RNA Club Individual talk Keeping developmental time with RNA 23.03.2017 Cambridge, Great Britain and Northern Ireland Grosshans Helge;
Seminar Individual talk Keeping developmental time with RNA 25.11.2016 IGBMC, Strasbourg, France Grosshans Helge;
The Complex Life of mRNA (EMBO Conference Series) Poster LIN41 silences mRNAs by two distinct and position-dependent mechanisms 05.10.2016 Heidelberg, Germany Aeschimann Florian;
EMBO | EMBL Symposium: The Complex Life of mRNA Poster Mechanistic and developmental functions of LIN41/TRIM71 as a post-transcriptional silencer in C. elegans 05.08.2016 Heidelberg, Germany Liu Jun;


Communication with the public

Communication Title Media Place Year
Media relations: print media, online media A key player in the maturation of sexual organs Press release, picked up by various (Technology Networks, The Medical News, Phys.org, EurekAlert International 2019
Media relations: print media, online media Ein wichtiger Akteur bei der Reifung der Geschlechtsorgane Media Release, picked up widely (Blick, SDA, BZBasel, Aargauer Zeitung,...) German-speaking Switzerland 2019
Media relations: print media, online media Cracking the RNA-binding code of a cell fate regulator Press release, picked up e.g. by www.phys.org International 2018
Media relations: print media, online media Cell fate regulation by LIN41 determined by binding location Press Release, picked up e.g. by https://phys.org/ International 2017
Talks/events/exhibitions Scientifica 2017 (Züricher Wissenschaftstage ETH Zürich und Universität Zürich German-speaking Switzerland 2017

Awards

Title Year
Titularprofessor (University of Basel) 2019
Ed Fischer Prize (best PhD Thesis of the year at FMI) 2017

Associated projects

Number Title Start Funding scheme
143313 Mechanisms and Developmental Functions of MicroRNA Turnover: The Exoribonuclease XRN2 01.02.2013 Project funding (Div. I-III)
188487 Mechanisms that time the onset of adulthood 01.11.2019 Project funding (Div. I-III)

Abstract

Faithful development of an organism requires precise regulation of cell fates in both space and time. Tight control of gene expression through both transcriptional and posttranscriptional mechanisms helps to achieve this. The Caenorhabditis elegans heterochronic pathway provides a prominent example of the importance of posttranscriptional mechanisms, as it relies extensively on mRNA silencing by microRNAs (miRNAs) to specify temporal cell fates during development. Most notably, the heterochronic let-7 miRNA terminates self-renewal activity in epidermal seam cells (a type of progenitor cell) and promotes vulval development. Loss of its activity is lethal as a consequence of, remarkably, dysregulation of only a single target, the TRIM-NHL (Tripartite Motif, NCL-1, HT2A2, and LIN-41 repeat domain) protein LIN-41/TRIM71. Indeed, let-7 and its regulation of LIN-41/TRIM71 appear to constitute an ancient control mechanism, conserved from invertebrates to vertebrates, for self-renewal, differentiation, and cell fate plasticity in diverse tissues. Nonetheless, little is known about the molecular function(s) and downstream effectors of LIN-41/TRIM71. We will use molecular genetic, biochemical and genomics approaches to study the mechanism of action and developmental roles of LIN-41 in C. elegans. We will test whether and to what extent regulation of different targets contributes to different developmental functions of LIN-41 in seam cells, vulva, and other tissues. Moreover, as preliminary data indicate LIN-41's mode of action to be target-dependent, we will seek to identify the elements of both the LIN-41 protein and its target RNAs that direct transcript degradation and translational repression, respectively. Given the conserved nature of the let-7 - LIN-41/TRIM71 regulatory module, we expect to gain new insights into a fundamental mechanism of posttranscriptional regulation of development.
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