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The long isoform of the C. elegans ELT-3 GATA factor can specify endoderm when overexpressed.

ABSTRACT: The C. elegans elt-3 gene encodes a GATA transcription factor that is expressed in the hypodermis and has roles in hypodermal specification and regulation of collagen and stress response genes. The gene encodes short and long isoforms, ELT-3A and ELT-3B respectively, that differ upstream of their DNA-binding domains. Previous work showed that ELT-3A can specify hypodermal cell fates when forcibly overexpressed throughout early embryos. We recently showed that the ELT-3B orthologue from the distantly related species C. angaria can specify endodermal fates when forcibly overexpressed in C. elegans. Here, we show that C. elegans ELT-3B can also specify endoderm.

SUBMITTER: Broitman-Maduro G

PROVIDER: S-EPMC9898813 | biostudies-literature | 2023

REPOSITORIES: biostudies-literature

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The long isoform of the <i>C. elegans</i> ELT-3 GATA factor can specify endoderm when overexpressed.

Broitman-Maduro Gina G Maduro Morris F MF

microPublication biology 20230121

The <i>C. elegans elt-3</i> gene encodes a GATA transcription factor that is expressed in the hypodermis and has roles in hypodermal specification and regulation of collagen and stress response genes. The gene encodes short and long isoforms, ELT-3A and ELT-3B respectively, that differ upstream of their DNA-binding domains. Previous work showed that ELT-3A can specify hypodermal cell fates when forcibly overexpressed throughout early embryos. We recently showed that the ELT-3B orthologue from th ...[more]

PMID: 36748041

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Project description:The two GATA transcription factors ELT-2 and ELT-7 function in the differentiation of the C. elegans intestine. ELT-2 loss causes lethality. ELT-7 loss causes no obvious phenotype but enhances the elt-2(-) intestinal phenotype. Thus, ELT-2 and ELT-7 appear partially redundant, with ELT-2 being more influential. To investigate the different regulatory roles of ELT-2 and ELT-7, we compared the transcriptional profiles of pure populations of wild-type, elt-2(-), elt-7(-), and elt-7(-); elt-2(-) double mutant L1-stage larvae. Consistent with the mutant phenotypes, loss of ELT-2 had a>25 fold greater influence on the number of significantly altered transcripts compared to the loss of ELT-7; nonetheless, the levels of numerous transcripts changed upon loss of ELT-7 in the elt-2(-) background. The quantitative responses of individual genes revealed a more complicated behaviour than simple redundancy/partial redundancy. In particular, genes expressed only in the intestine showed three distinguishable classes of response in the different mutant backgrounds. One class of genes responded as if ELT-2 is the major transcriptional activator and ELT-7 provides variable compensatory input. For a second class, transcript levels increased upon loss of ELT-2 but decreased upon further loss of ELT-7, suggesting that ELT-7 actually overcompensates for the loss of ELT-2. For a third class, transcript levels also increased upon loss of ELT-2 but remained elevated upon further loss of ELT-7, suggesting overcompensation by some other intestinal transcription factor(s). In spite of its minor loss-of-function phenotype and its limited sequence similarity to ELT-2, ELT-7 expressed under control of the elt-2 promoter is able to rescue elt-2(-) lethality. Indeed, appropriately expressed ELT-7, like appropriately expressed ELT-2, is able to replace all other core GATA factors in the C. elegans endodermal pathway. Overall, this study focuses attention on the quantitative intricacies behind apparent redundancy or partial redundancy of two related transcription factors.

Project description:The ELT-2 GATA factor normally functions in differentiation of the C. elegans endoderm, downstream of endoderm specification. We have previously shown that, if ELT-2 is expressed sufficiently early, it is also able to specify the endoderm and to replace all other members of the core GATA-factor transcriptional cascade (END-1, END-3, ELT-7). However, such rescue requires multiple copies (and presumably overexpression) of the end-1p::elt-2 cDNA transgene; a single copy of the transgene does not rescue. We have made this observation the basis of a genetic screen to search for genetic modifiers that allow a single copy of the end-1p::elt-2 cDNA transgene to rescue the lethality of the end-1 end-3 double mutant. We performed this screen on a strain that has a single copy insertion of the transgene in an end-1 end-3 background. These animals are kept alive by virtue of an extrachromosomal array containing multiple copies of the rescuing transgene; the extrachromosomal array also contains a toxin under heat shock control to counterselect for mutagenized survivors that have been able to lose the rescuing array. A screen of ?14,000 mutagenized haploid genomes produced 17 independent surviving strains. Whole genome sequencing was performed to identify genes that incurred independent mutations in more than one surviving strain. The C. elegans gene tasp-1 was mutated in four independent strains. tasp-1 encodes the C. elegans homolog of Taspase, a threonine-aspartic acid protease that has been found, in both mammals and insects, to cleave several proteins involved in transcription, in particular MLL1/trithorax and TFIIA. A second gene, pqn-82, was mutated in two independent strains and encodes a glutamine-asparagine rich protein. tasp-1 and pqn-82 were verified as loss-of-function modifiers of the end-1p::elt-2 transgene by RNAi and by CRISPR/Cas9-induced mutations. In both cases, gene loss leads to modest increases in the level of ELT-2 protein in the early endoderm although ELT-2 levels do not strictly correlate with rescue. We suggest that tasp-1 and pqn-82 represent a class of genes acting in the early embryo to modulate levels of critical transcription factors or to modulate the responsiveness of critical target genes. The screen's design, rescuing lethality with an extrachromosomal transgene followed by counterselection, has a background survival rate of <10-4 without mutagenesis and should be readily adapted to the general problem of identifying suppressors of C. elegans lethal mutations.

Dataset Information

The long isoform of the C. elegans ELT-3 GATA factor can specify endoderm when overexpressed.

Publications

The long isoform of the <i>C. elegans</i> ELT-3 GATA factor can specify endoderm when overexpressed.

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