MEX-5 asymmetry in one-cell C. elegans embryos requires PAR-4- and PAR-1-dependent phosphorylation.

Anteroposterior polarity in early C. elegans embryos is required for the specification of somatic and germline lineages, and is initiated by a sperm-induced reorganization of the cortical cytoskeleton and PAR polarity proteins. Through mechanisms that are not understood, the kinases PAR-1 and PAR-4, and other PAR proteins cause the cytoplasmic zinc finger protein MEX-5 to accumulate asymmetrically in the anterior half of the one-cell embryo. We show that MEX-5 asymmetry requires neither vectorial transport to the anterior, nor protein degradation in the posterior. MEX-5 has a restricted mobility before fertilization and in the anterior of one-cell embryos. However, MEX-5 mobility in the posterior increases as asymmetry develops, presumably allowing accumulation in the anterior. The MEX-5 zinc fingers and a small, C-terminal domain are essential for asymmetry; the zinc fingers restrict MEX-5 mobility, and the C-terminal domain is required for the increase in posterior mobility. We show that a crucial residue in the C-terminus, Ser 458, is phosphorylated in vivo. PAR-1 and PAR-4 kinase activities are required for the phosphorylation of S458, providing a link between PAR polarity proteins and the cytoplasmic asymmetry of MEX-5.

EEL-1, a Hect E3 ubiquitin ligase, controls asymmetry and persistence of the SKN-1 transcription factor in the early C. elegans embryo.

During early divisions of the C. elegans embryo, many maternally supplied determinants accumulate asymmetrically, and this asymmetry is crucial for proper cell fate specification. SKN-1, a transcription factor whose message is maternally supplied to the embryo, specifies the mesendodermal cell fate. In the 2-cell embryo, SKN-1 is expressed at a higher level in the posterior cell. This asymmetry becomes more pronounced at the 4-cell stage, when SKN-1 is high in the posterior cell's daughters and low in the daughters of the anterior blastomere. To date, the direct mechanisms that control SKN-1 distribution remain unknown. In this report, we identify eel-1, which encodes a putative Hect E3 ubiquitin ligase that shares several domains of similarity to the mammalian E3 ligase Mule. EEL-1 binds SKN-1 and appears to target SKN-1 for degradation. EEL-1 has two functions in regulating SKN-1 during early embryogenesis. First, eel-1 promotes the spatial asymmetry of SKN-1 accumulation at the 2- and 4-cell stages. Second, eel-1 acts in all cells to downregulate SKN-1 from the 12- to the 28-cell stage. Although loss of eel-1 alone causes a reduction in SKN-1 asymmetry at the 2-cell stage, the function of eel-1 in both the spatial and temporal regulation of SKN-1 is redundant with the activities of other genes. These data strongly suggest that multiple, functionally redundant pathways cooperate to ensure precise control of SKN-1 asymmetry and persistence in the early embryo.

Reduced dosage of pos-1 suppresses Mex mutants and reveals complex interactions among CCCH zinc-finger proteins during Caenorhabditis elegans embryogenesis.

Cell fate specification in the early C. elegans embryo requires the activity of a family of proteins with CCCH zinc-finger motifs. Two members of the family, MEX-5 and MEX-6, are enriched in the anterior of the early embryo where they inhibit the accumulation of posterior proteins. Embryos from mex-5 single-mutant mothers are inviable due to the misexpression of SKN-1, a transcription factor that can specify mesoderm and endoderm. The aberrant expression of SKN-1 causes a loss of hypodermal and neuronal tissue and an excess of pharyngeal muscle, a Mex phenotype (muscle excess). POS-1, a third protein with CCCH motifs, is concentrated in the posterior of the embryo where it restricts the expression of at least one protein to the anterior. We discovered that reducing the dosage of pos-1(+) can suppress the Mex phenotype of mex-5(-) embryos and that POS-1 binds the 3'-UTR of mex-6. We propose that the suppression of the Mex phenotype by reducing pos-1(+) is due to decreased repression of mex-6 translation. Our detailed analyses of these protein functions reveal complex interactions among the CCCH finger proteins and suggest that their complementary expression patterns might be refined by antagonistic interactions among them.