We have been studying genes that are involved in the development of the HSN neurons. One of these genes,
ham-1 IV, appears to be involved not only in HSN development but also in the development of the phasmid neuron PHB, the sister cell of the HSN. The HSN/PHB phenotypes led us to propose a model in which the distribution of developmental potential between the HSN and PHB is abnormal in
ham-1 animals (Desai et al., 1988). In order to understand how the
ham-1 gene product functions in HSN and PHB development, we have begun a molecular analysis of this gene. Using Bristol (N2)/Bergerac (N62) RFLPs as genetic markers, we mapped the
ham-1 gene to the contig containing
unc-31. A more detailed analysis mapped
ham-1 between RFLPs recognized by the cosmids C08G1 and C13H6; of the 74 recombinants analyzed,
ham-1 was not separated from the RFLP recognized by C03F9 (C08G1 and C13H6 overlap C03F9). To see if we could rescue the
ham-1 defects with genomic DNA from this region, we coinjected into
ham-1 animals the
rol-6 plasmid pRF4 together with cosmids that mapped near
ham-1. From the progeny of the injected animals, we established roller lines and scored them for two
ham-1 phenotypes, egg laying and the ability of the phasmids to fill with fluorescent dyes. Two cosmids, C03F9 and C13H6, rescued
ham-1 animals for both egg-laying and phasmid loading defects, indicating that both C03F9 and C13H6 contain the
ham-1 gene. In additional experiments we have rescued the
ham-1 defects with a 23 kbp fragment of C03F9 and partially rescued the
ham-1 defects with a 12 kbp fragment. On Northern blots of poly(A)+ RNA using probes spanning C03F9, we detected a 1.8 kb RNA that we believe is the
ham-1 mRNA for two reasons. First, it is the only transcript that is detected on Northern blots when the 12 kbp DNA that partially rescues
ham-1 is used as a probe. Second, it is the only transcript recognized by the probes spanning C03F9 that is significantly reduced (>95%) in RNA from the mutant
ham-1(
n1438). Genomic DNA from
ham-1(
n1438) contains a small deletion in this region. In additional Northern blot experiments, we have shown that the
ham-1 mRNA is enriched in poly(A)+ RNA prepared from embryos. We screened 100,000 plaques from Stuart Kim's cDNA library with the 23 kbp fragment that rescues
ham-1 and isolated four cDNAs. One appears to be a
ham-1 cDNA because it comes from the 12 kbp region that partially rescues
ham-1 and it recognizes the 1.8 kb
ham-1 mRNA on Northern blots. Since the cDNA is 1.8 kbp, it contains most of the
ham-1 RNA. Sequence analysis of this cDNA revealed a single long open reading frame that could encode a protein of 414 amino acids. The putative initiator methionine is preceded by three in-frame stop codons indicating that we have defined the entire
ham-1 coding region. This protein has no significant sequence similarity to any protein in the gene bank (TRANSGEN). Add
ham-1 to the scores of novel developmentally important genes defined by C. elegans' genetics.