Med-type GATA factors and the evolution of mesendoderm specification in nematodes

Abstract

In the nematode, C. elegans, the divergent GATA-type transcription factors MED-1 and MED-2 are encoded by an unlinked, redundant pair of intronless genes. The med-1,2 genes are among the first to be activated in the embryo and are critical for the specification of the 7-cell stage MS (mesoderm) and E (endoderm) precursor cells. We have previously shown that the binding site recognized by MED-1 is a noncanonical RAGTATAC site that is not expected from the resemblance of its single C4-type zinc finger to those of other known GATA factors, which recognize the consensus HGATAR. To date, no MED-like zinc fingers have been described outside of C. elegans. In order to understand the evolution of these transcription factors, and the evolution of gene networks that specify early cell fates in Caenorhabditis, we have identified med sequence homologs in the related nematodes C. briggsae and C. remanei. While C. briggsae encodes two med-like genes similar to C. elegans, we find evidence for seven distinct med-like genes in C. remanei. Somewhat unexpectedly, the coding regions of all med genes appear to lack introns. We report that the med homologs have similar expression in their respective species. We further show that the C. briggsae homologs, and at least five of the seven C. remanei homologs, can fully complement the embryonic lethal phenotype of a C. elegans med-1,2(-) strain. We conclude that Med function and expression have been conserved over tens of millions of years of evolution, and that there may be a mechanism that selects against the acquisition of introns in these genes.

The NK-2 class homeodomain factor CEH-51 and the T-box factor TBX-35 have overlapping function inC. elegansmesoderm development

“…Embryos were stained as described (Coroian et al, 2005). For pal-1 staining of med-1,2 and ceh-51; tbx-35 embryos, a mixture of rescued and nonrescued embryos were stained, and the number of mutants was estimated from the array transmission frequency.…”

“…In skn-1 mutants, mis-specified MS and E cells adopt the fate of the mesectodermal precursor C (Bowerman et al, 1992). In EMS (the mother of MS and E), SKN-1 activates the zygotic med-1 med-2 (med-1,2) divergent GATA factor gene pair (Coroian et al, 2005;Maduro et al, 2001). Loss of med-1,2 has a similar effect on MS specification as loss of skn-1, but a much weaker effect on E specification owing to parallel contributions to endoderm from SKN-1 and other factors (Goszczynski and McGhee, 2005;Maduro et al, 2005a;Maduro et al, 2001).…”

Neither Maternal nor Zygotic med-1/med-2 Genes Play a Major Role in Specifying the Caenorhabditis elegans Endoderm

“…1 As reported in detail elsewhere , we had only limited success producing med-type-arrested embryos by either of these two protocols. With the first protocol, we were able to produce fewer than one gut-granule-negative embryo for every two injected hermaphrodites, which is only marginally higher than that produced by injection of nonspecific control RNA (see also Coroian et al 2006). In our hands, the second protocol produces either sterile mothers or embryos that arrest prior to the stage where they would ordinarily express gut granules.…”

“…In the first protocol, embryos were collected 7-9 hr following injection of concentrated doublestranded RNA (dsRNA) into the maternal gonad; the rationale for such an early and limited observation period was to more effectively target the transient expression of the med-1/2 genes in the early embryo (Maduro et al 2001;Coroian et al 2006). In the more standard protocol, the RNAi effect is found to be maximally effective from 1 to 3 days following injection (Fire et al 1998;Zipperlen et al 2001;Goszczynski and McGhee 2005;Ahringer 2006); however, these more usual methods of administering RNAi show no effect with the med-1/2 genes (Kamath et al 2003;Goszczynski and McGhee 2005;Sonnichsen et al 2005).…”

“…A second method of administering med-1/2 RNAi (Maduro et al 2001) was by means of a transgenic heat-inducible promoter driving expression of double-stranded med RNA (Tavernarakis et al 2000). Together, these two protocols were reported to produce a small number of embryos that arrested with a distinctive phenotype (approximately twofold elongation with absence of posterior pharynx); $50% of these arrested embryos failed to show the standard marker for endoderm specification, the presence of birefringent gut granules (Maduro et al 2001;Coroian et al 2006). 1 As reported in detail elsewhere , we had only limited success producing med-type-arrested embryos by either of these two protocols.…”