Worm Breeder's Gazette 11(5): 31

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 Axonal Outgrowth Gene unc-76 Encodes an Unfamiliar Protein

Laird Bloom and Bob Horvitz

The genes unc-34 and unc-76 are required for normal outgrowth of 
many neuronal processes.  Mutations in either gene cause several 
classes of neurons to terminate outgrowth prematurely (HSN motor 
neurons (1), PHA, PHB, PDE, and amphidial sensory neurons (2)) or to 
follow abnormal paths (VD and DD motor neurons (3), AVM 
mechanoreceptors (4)).  Our studies have led us to believe that these 
genes function generally in axonal guidance and organization of axon 
We previously reported that several cosmids rescue the unc-76 
e in germline transformation experiments (WBG 
11(3): 26).  Two rescuing cosmids, C56C4 and C08C1, have 27 kb in 
common.  A 15 kb subclone from this region of C56C4 gives full 
rescuing activity, and smaller subclones of 8.5, 7.0, 6.2, and 5.5 kb 
rescue the Unc phenotype of young larvae but give weaker rescue in 
older animals.
We screened 220,000 plaques from Stuart Kim's mixed-stage cDNA 
library using the 8.5 kb subclone as a probe and identified five cDNA 
clones.  Two of these clones, 1.8 and 2.4 kb in size, hybridize to the 
5.5 kb rescuing region.  Both of these clones hybridize to a 1.8 kb 
RNA band in Northern blots of embryonic poly-A(+) RNA.  We believe 
that this RNA represents the unc-76 transcript because the amount of 
this RNA in unc-76(e911) embryos is much lower than that in N2 embryos.
unc-76(e911) is weakly suppressed by smg suppressors, which have 
been shown to elevate the level of message in certain unc-54 mutants 
in which the amount of message is reduced (5).  However, the amount of 
the 1.8 kb message in smg-1(e1228) him-2(e1065); 11)
embryos is only slightly higher than that in unc-76(e911) embryos.
Sequence analysis of the 1.8 kb cDNA identified a single long open 
reading frame that could encode a 379-amino acid polypeptide.  A 
search of the Genbank database using the FASTA program revealed no 
significant similarities to known proteins.  Neither cDNA clone 
appears to contain the 5' end of the gene, but the 2.4 kb cDNA clone 
includes an additional 130 bp at the 5' end of the open reading frame, 
extending it to 422 amino acids.  Comparison of the restriction maps 
of the two cDNA clones suggests that the 2.4 kb clone contains about 0.
5 kb of DNA 3' to the sequences contained in the 1.8 kb cDNA, but we 
do not yet know whether this 0.5 kb of additional DNA represents a 
cloning artifact or an alternative splicing product with additional 3'