Worm Breeder's Gazette 11(4): 38

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.

The ham-1 Gene Encodes an Unfamiliar Protein

Gian Garriga and Bob Horvitz

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.