Worm Breeder's Gazette 13(1): 47 (October 1, 1993)
These abstracts should not be cited in bibliographies. Material contained herein should be treated as personal communication and should be cited as such only with the consent of the author.
During the development of the C. elegans embryo, many cells undergo long range migrations. For example, the HSNs, a pair of serotonergic motor neurons, migrate anteriorly from their birthplace in the tail to positions just lateral to the gonad primordium. We are studying the development of the HSNs in an attempt to understand the molecular mechanisms involved in cell migration. Mutation, in 19 genes affect HSN migration. Of these, mutations in the gene egl-43 result in the most severe HSN migration defect. In egl-43 mutants, the HSNs traverse on average only 5-10% of the distance migrated by HSNs in wild-type hermaphrodites. egl-43 encodes two putative zinc-finger proteins with sequence similarity to a set of transcription factors first defined by the Xenopus transcription factor TFIIIA. This has led us to propose a model where egl-43 acts to transcriptionally regulate genes that are more directly involved in cell migration. We might expect egl-43 to function in the HSNs. Alternatively, egl-43 could function in surrounding cells to provide a signal for HSN migration.
In order to test these hypotheses, we were interested in determining the expression pattern of egl-43 .Polyclonal antibodies against three regions of egl-43 were generated by injecting mice with fusion proteins. Two of the three antibodies were affinity purified and used to stain animals that had been fixed as described in Finney and Ruvkin, Cell 63:895-905.
Consistent with a role for egl-43 as a transcription factor, egl-43 protein is localized to the nuclei of certain cells beginning at about the 64 cell stage in embryos. To facilitate the identification of cells expressing egl-43 ,animals were double stained with antibodies recognizing the egl-43 and unc-86 hasmidneurons, the two PVD neurons, two cells nearby the PVD neurons, and additional cells in the head. However, egl-43 expression is never observed in the HSNs at stages of development where we are able to identify the HSNs.
The presence of egl-43 protein in the phasmid neurons but not in the HSNs is particularly interesting. It suggests that the primary defect in egl-43 mutants might be in the phasmid neurons and that the HSN migration defect might be secondary, perhaps resulting from a failure of the hasmid neurons, the two PVD neurons, two cells nearby the PVD neurons, and additional cells in the head. However, egl-43 expression is never observed in the HSNs at stages of development where we are able to identify the HSNs.
The presence of egl-43 protein in the phasmid neurons but not in the HSNs is particularly interesting. It suggests that the primary defect in egl-43 mutants might be in the phasmid neurons and that the HSN migration defect might be secondary, perhaps resulting from a failure of the phasmid neurons to provide a migration cue to the HSNs. Several pieces of data are consistent with the phasmid neurons being affected in egl-43 mutants. Dye-loading experiments using carbocyanine dyes (diI and diO) and electron microscopy demonstrated that the phasmid neurons are variably defective in egl-43 mutants. In addition, a comparison of N2 and the egl-43 ( n1 O79)mutant stained with the egl-43 antiserum indicates that the phasmid neurons are abnormal in egl-43 ( n1 O79)mutants. In wild-type L1 larvae, both PHA and PHB stained in 96% of the sides examined, whereas only one of the phasmid neurons stained in the other 4% of the sides (n=250). In egl-43 ( n1 O79) L1 larvae, both PHA and PHB stained in 9.6% of the sides examined, only one of the phasmid neurons stained in 66.5% of the sides, and neither phasmid neuron stained in 23.9% of the sides (n=230). We have not determined if the phasmid neuron stained in egl-43 mutants is consistently PHA or PHB. We have also not determined whether any of the cells visible in the head of wild-type animals are defective in egl-43 mutants.
In light of these results, egl-43 might regulate genes necessary for phasmid neuron development or for the transmission of a signal from the phasmid neurons to the HSNs. We are currently confirming the identifications of those cells expressing egl-43 and are considering laser ablation and mosaic experiments that would directly address the role of the phasmid neurons and additional egl-43 expressing cells in HSN migration.