daf-42 is an evolutionarily young gene essential for dauer development in Caenorhabditis elegans

Background:

Dauer diapause – a state of hypobiosis – is widely conserved in many nematodes and appears to have driven the evolution of parasitism. For over five decades, free-living C. elegans has provided a convenient model to study the molecular mechanism of dauer formation. Dauers exhibit a distinct morphology which includes a specialized thickened cuticle with alae. Their body is constricted, and they develop a buccal plug that seals the mouth – all to promote survival in harsh environments (Cassada & Russell, 1975; Fielenbach & Antebi, 2008).

In C. elegans, the dauer developmental decision occurs at the late L1 stage though the commitment to dauer formation does not occur until mid-late L2d stage (Figure 1). Dauer entry is influenced by environmental factors (food availability, temperature, and pheromone) and is regulated by multiple signaling pathways, including the conversed insulin signaling pathway, TGFβ, and others. Mutants that show a dysfunctional dauer/adult developmental switch can be broadly divided into two classes: dauer-constitutive (Daf-c) and dauer-defective (Daf-d). While Daf-c mutants always enter dauer diapause, Daf-d animals never form dauer even in the presence of environmental triggers (Fielenbach & Antebi, 2008).

In this preprint, Lim and colleagues report a novel regulator of dauer development – DAF-42. Unlike the “classical” Daf-c and Daf-d that regulate the dauer entry decision, DAF-42 promotes L2d-to-dauer transition. Interestingly, despite the essential role of DAF-42 in dauer development, it is poorly conserved outside of the Caenorhabditis genus, suggesting rapid evolution of the dauer gene regulatory network.

Figure 1: C. elegans life cycle. C. elegans first stage (L1) larvae grows into reproductive adults in ~3 days at 20^OC. In unfavorable growth conditions, the L1 animal can develop to form dauer, which can live several months without food. The dauer can resume reproductive development when the environment recovers.

Major findings:

• daf-42 is a novel gene required for dauer entry.

By combining a screen for C. elegans mutants defective in dauer formation with genetic mapping-by-sequencing, the authors found that mutants lacking daf-42 failed to develop into dauer and would die when grown under dauer-inducing conditions. Next, the authors introduced the daf-42 mutation into worms carrying daf-2(e1370), a temperature sensitive Daf-c mutation, and could show that daf-2(-); daf-42(-) double mutants arrest at the L2d stage, while daf-2(-) single mutants robustly developed into dauers at 25^oC. daf-2(-); daf-42(-) double mutants showed a degraded pharynx and molting defects. Moreover, dauer alae and cuticles were not properly formed in daf-2(-); daf-42(-) animals, indicating that daf-2(-); daf-42(-) animals can initiate the dauer formation program but fail to complete the developmental transition. Supporting this notion, removing DAF-7 (TGFβ ligand) or DAF-9 (cytochrome p450 enzyme in the steroid hormone pathway), both of which regulate the dauer entry decision, failed to rescue daf-42(-) lethality as DAF-42 functions downstream of the decision point influenced by harsh environmental conditions.

• DAF-42 is an unstructured secreted protein that promotes L2d-to-dauer transition.

The authors reported that DAF-42 is a large protein with 17 isoforms. It is largely disordered and contains a signal peptide at the N terminus.

By examining transcriptomic data and transgenic reporters, the authors found that daf-42 is expressed in the seam cells (specialized hypodermal cells) during dauer entry. Indeed, expressing daf-42 under the control of a seam cell promotor could partially rescue the lethality of daf-2(-); daf-42(-) double mutants. The authors could show that DAF-42 is secreted from the seam cell to the peripheral hypodermis and cuticle during L2d stage, perhaps to promote molting during the L2d-to-dauer transition.

Next, the authors examined the transcriptomic differences between daf-2(-) single and daf-2(-); daf-42(-) double mutants and found that daf-2(-); daf-42(-) worms showed significant changes in gene expression during L2d, but not at an earlier point, supporting the model in which DAF-42 acts after dauer commitment in L2d. Intriguingly, many genes upregulated in daf-2(-); daf-42(-) are related to immune system and defense response.

• daf-42 is a newly evolved gene in the Caenorhabditis

It is clear that DAF-42 plays an essential role in dauer development. Surprisingly, however, DAF-42 homologs are not found in nematodes outside of the Caenorhabditis genus. Within Caenorhabditis, the DAF-42 protein sequences exhibit substantial variation. These results indicate that the gene regulatory network for dauer development is highly plastic and a young gene has quickly evolved to gain essential developmental functions.

What I like about this preprint:

This preprint identifies an unusual daf gene that promotes dauer development. The data presented is solid. Most surprisingly, daf-42 appears to be evolutionary young and to have quickly evolved to adopt an essential developmental role. This study has shed light on the evolution of the nematode dauer signalling pathway and provides an interesting paradigm to further study the evolutionary trajectory of young genes.

Questions for the authors:

• For the translational reporter, is it possible that not all isoforms are tagged, and daf-42 expression is not fully captured?
• Have you looked closer at the fate of DAF-42 in the target tissues? Does it, for example, get taken up by the hypodermal cells?
• It appears that DAF-42 may play an important role in molting as evident by the mutant phenotype and daf-42 expression pattern. Did you observe changes in the expression of genes regulating molting in the mutant?

References:

Cassada, R. C., & Russell, R. L. (1975). The dauerlarva, a post-embryonic developmental variant of the nematode Caenorhabditis elegans. Developmental Biology, 46(2), 326–342. https://doi.org/10.1016/0012-1606(75)90109-8

Fielenbach, N., & Antebi, A. (2008). C. elegans dauer formation and the molecular basis of plasticity. Genes & Development, 22(16), 2149–2165. https://doi.org/10.1101/gad.1701508

 

Tags: dauer, developmental plasticity, diapause, molting

doi: https://doi.org/10.1242/prelights.34630

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