The ELT-3 GATA factor specifies endoderm in Caenorhabditis angaria in an ancestral gene network

Gina Broitman-Maduro, Morris F. Maduro

Preprint posted on May 26, 2022

How many ways to make a worm gut? Extensive rewiring of the gene network underlying gut development in Caenorhabditis

Selected by Chee Kiang Ewe


Development is driven by gene regulatory networks (GRNs) that are composed of complex interactions between many transcription factors. The nematode C. elegans has been a great system to study the principals underlying developmental GRN owing to its defined cell lineage and the extensive genetic tools available for manipulating the GRN. Furthermore, comparative studies involving different nematode species spanning the Caenorhabditis genus (Figure 1) have provided insights into the evolutionary variation in developmental mechanisms, and how GRNs might be modified during evolution (Ewe et al., 2020).

Figure 1: The phylogenetic relationships of the Caenorhabditis nematodes. MEDs, ENDs, and ELT-7 originate at the base of the Elegans Supergroup (adapted from Figure 1; Broitman-Maduro & Maduro, 2022).

In C. elegans, the endoderm comprises the mid-gut. In the early embryo, the endoderm GRN is initiated by SKN-1, which activates a gene cascade driven by six GATA transcription factors – two MEDs, two ENDs, and two ELTs. (Figure 2). Additionally, the Wnt effector POP-1 activates endoderm specification by acting on END-1 and -3. Interestingly, the orthologues of the MEDs and ENDs, as well as ELT-7, are missing in nematodes outside of the Elegans Supergroup, including C. angaria and C. portoensis (Figure 1) (Maduro, 2020), raising the question of how gut is specified in their absence.

In this preprint, the authors found that ELT-3, a GATA factor that is mostly expressed in the hypodermal cells in C. elegans, is present in early endodermal progenitors and drives gut specification in C. angaria. This study demonstrates an extensive rewiring of the endoderm GRN within the Caenorhabditis genus (Figure 2).

Figure 2: The evolution of the endoderm GRN in nematodes. POP-1 activates ELT-3, which in turns activates ELT-2 in C. angaria. In the derived GRN in C. elegans, MEDs and ENDs, instead of ELT-3, drive endoderm specification. This eventually leads to the activation of ELT-7 and -2, which lock down the differentiated endodermal cell fate.

Major findings:

1) Wnt effector POP-1 is required for embryonic gut development in C. angaria

To test the roles of SKN-1 and POP-1 orthologues in endoderm development in C. angaria, the authors knocked down Can-skn-1 and Can-pop-1 using RNAi. While knocking down Can-skn-1 did not cause obvious developmental defects, embryos lacking Can-pop-1 underwent developmental arrest and failed to development a gut.

2) GATA factor ELT-2 plays conserved role in endoderm differentiation

To examine the role of C. angaria ELT-2 in endoderm development, the authors cloned Can-elt-2 and introduced the transgene into C. elegans. They found that Can-elt-2, like Cel-elt-2, was expressed exclusively in the endodermal lineage. In C. elegans, eliminating both elt-2 and elt-7 causes larval arrest and a severe block to endoderm differentiation (Sommermann et al., 2010). The authors showed that the expression of Can-elt-2 transgene strongly rescued larval lethality in C. elegans double mutant lacking elt-2 and elt-7. Importantly, knocking down Can-elt-2 in C. angaria caused penetrant larval lethality and gut differentiation defects. Together, these results demonstrate the functional conservation of ELT-2 in endoderm differentiation.

3) ELT-3 drives endoderm specification in C. angaria

The authors showed that the expression of Can-elt-2 in C. elegans required END-1 and -3, the two endoderm-specifying GATA factors, as knocking out end-1/3 abrogated Can-elt-2 transgene expression; however, C. angaria does not contain END-1 and -3 orthologues. To identify the upstream activator(s) of Can-elt-2, the authors performed smFISH to examine the expression of other GATA-encoding genes (elt-1, elt-3, and elt-5) in C. angaria embryos. Interestingly, the authors found that the transcripts of elt-3, but not elt-1 and -5, were found in the early endodermal lineage, and the expression of Can-elt-3 depended on Can-POP-1, but not Can-SKN-1.

To examine the role of elt-3 in endoderm specification, the authors overexpressed Can-elt-3 (under the control of a heat-shock promotor) in C. elegans and found a widespread activation Can-elt-2 and ectopic gut development in the arrested embryos. Furthermore, knocked down elt-3 in C. angaria by RNAi resulted in development arrest and a severe block to endoderm development, showing the function of Can-ELT-3 as a potent driver of endoderm specification, akin to END-1 and -3 in C. elegans.

4) The evolution of the endoderm gene regulatory network in nematodes

Lastly, the authors found elt-3 and elt-2 to be expressed in the endodermal lineage of C. portoensis, another species outside of the Elegans Supergroup, suggesting that ELT-3 -> ELT-2 interaction is conserved in the ancestral endoderm GRN, prior to the gene duplication events that gave rise to meds, ends, and elt-7 (Figure 2). The intercalation of the redundant GATA factors in the endoderm GRN may ensure developmental robustness during rapid embryogenesis in C. elegans.

What I liked about this preprint: This work provides an important insight into the dynamics of developmental system drift in the endoderm GRN in nematodes – an excellent paradigm to study the molecular mechanisms of developmental robustness and evolutionary plasticity.

Question for the authors:

The genetic redundancy in C. elegans may indeed promote developmental robustness; the simplicity of the ancestral gene network may therefore result in increased transcriptional noise. Have you noted obvious variability in elt-2 onset timing and/or expression level in C. angaria?


Ewe, C. K., Torres Cleuren, Y. N., & Rothman, J. H. (2020). Evolution and developmental system drift in the endoderm gene regulatory network of Caenorhabditis and other nematodes. Frontiers in Cell and Developmental Biology, 8, 170.

Maduro, M. F. (2020). Evolutionary dynamics of the Skn-1/Med/end-1,3 regulatory gene cascade in Caenorhabditis endoderm specification. G3: Genes, Genomes, Genetics, 10(1), 333–356.

Sommermann, E. M., Strohmaier, K. R., Maduro, M. F., & Rothman, J. H. (2010). Endoderm development in Caenorhabditis elegans: The synergistic action of ELT-2 and -7 mediates the specification→differentiation transition. Developmental Biology, 347(1), 154–166.


Tags: embryogenesis, endoderm, gata, nematodes

Posted on: 11th June 2022


Read preprint (No Ratings Yet)

Author's response

Morris F. Maduro shared

As far as variability in elt-2 onset in C. angaria, we have not looked quantitatively yet, and of course there are likely to be other regulatory inputs that we do not know about, so although the network is simpler as far as GATA factors go, there may be other non-GATA factors (speculated by Rifkin’s paper the week before ours also in bioRxiv) that could be providing robustness. We also working on making a mutant in Can-elt-3, and to move the network into C. elegans, both of which are not proving straightforward yet.

Have your say

Your email address will not be published. Required fields are marked *

This site uses Akismet to reduce spam. Learn how your comment data is processed.

Sign up to customise the site to your preferences and to receive alerts

Register here

preLists in the developmental biology category:

2nd Conference of the Visegrád Group Society for Developmental Biology

Preprints from the 2nd Conference of the Visegrád Group Society for Developmental Biology (2-5 September, 2021, Szeged, Hungary)


List by Nándor Lipták


The advances in fibroblast biology preList explores the recent discoveries and preprints of the fibroblast world. Get ready to immerse yourself with this list created for fibroblasts aficionados and lovers, and beyond. Here, my goal is to include preprints of fibroblast biology, heterogeneity, fate, extracellular matrix, behavior, topography, single-cell atlases, spatial transcriptomics, and their matrix!


List by Osvaldo Contreras

EMBL Synthetic Morphogenesis: From Gene Circuits to Tissue Architecture (2021)

A list of preprints mentioned at the #EESmorphoG virtual meeting in 2021.


List by Alex Eve

EMBL Conference: From functional genomics to systems biology

Preprints presented at the virtual EMBL conference "from functional genomics and systems biology", 16-19 November 2020


List by Jesus Victorino

Single Cell Biology 2020

A list of preprints mentioned at the Wellcome Genome Campus Single Cell Biology 2020 meeting.


List by Alex Eve

Society for Developmental Biology 79th Annual Meeting

Preprints at SDB 2020


List by Irepan Salvador-Martinez, Martin Estermann

FENS 2020

A collection of preprints presented during the virtual meeting of the Federation of European Neuroscience Societies (FENS) in 2020


List by Ana Dorrego-Rivas

Planar Cell Polarity – PCP

This preList contains preprints about the latest findings on Planar Cell Polarity (PCP) in various model organisms at the molecular, cellular and tissue levels.


List by Ana Dorrego-Rivas

Cell Polarity

Recent research from the field of cell polarity is summarized in this list of preprints. It comprises of studies focusing on various forms of cell polarity ranging from epithelial polarity, planar cell polarity to front-to-rear polarity.


List by Yamini Ravichandran

TAGC 2020

Preprints recently presented at the virtual Allied Genetics Conference, April 22-26, 2020. #TAGC20


List by Maiko Kitaoka et al.

3D Gastruloids

A curated list of preprints related to Gastruloids (in vitro models of early development obtained by 3D aggregation of embryonic cells). Updated until July 2021.


List by Paul Gerald L. Sanchez and Stefano Vianello

ASCB EMBO Annual Meeting 2019

A collection of preprints presented at the 2019 ASCB EMBO Meeting in Washington, DC (December 7-11)


List by Madhuja Samaddar et al.

EDBC Alicante 2019

Preprints presented at the European Developmental Biology Congress (EDBC) in Alicante, October 23-26 2019.


List by Sergio Menchero et al.

EMBL Seeing is Believing – Imaging the Molecular Processes of Life

Preprints discussed at the 2019 edition of Seeing is Believing, at EMBL Heidelberg from the 9th-12th October 2019


List by Dey Lab

SDB 78th Annual Meeting 2019

A curation of the preprints presented at the SDB meeting in Boston, July 26-30 2019. The preList will be updated throughout the duration of the meeting.


List by Alex Eve

Lung Disease and Regeneration

This preprint list compiles highlights from the field of lung biology.


List by Rob Hynds

Young Embryologist Network Conference 2019

Preprints presented at the Young Embryologist Network 2019 conference, 13 May, The Francis Crick Institute, London


List by Alex Eve

Pattern formation during development

The aim of this preList is to integrate results about the mechanisms that govern patterning during development, from genes implicated in the processes to theoritical models of pattern formation in nature.


List by Alexa Sadier

BSCB/BSDB Annual Meeting 2019

Preprints presented at the BSCB/BSDB Annual Meeting 2019


List by Dey Lab

Zebrafish immunology

A compilation of cutting-edge research that uses the zebrafish as a model system to elucidate novel immunological mechanisms in health and disease.


List by Shikha Nayar

Also in the evolutionary biology category: