Worm Breeder's Gazette 14(1): 51 (October 1, 1995)
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.
Department of Biology, Faculty of Science, Kyushu University, Hakozaki, Higashi-ku, Fukuoka 812-81, Japan
unc-14 and unc-51 mutants have abnormalities in the axonal
formation of many neurons. They also have similar defects in membrane
structures of axons. We previously reported that the unc-51 gene
encodes a novel serine / threonine kinase, and that a lacZ fusion gene
was expressed in many neurons. To analyze the function of this gene, we
screened UNC-51 binding proteins using an yeast two-hybrid system.
About 5 x 100000 clones of a C. elegans cDNA library ( kindly
gifted by R. Barstead ) was screened, and 16 positive clones were
obtained. By homology analysis, one of them turned out to be an unc-51
cDNA encoding a C-terminal region. Four of them were overlapping with
cDNA clones isolated by Yuji Kohara, which hybridized with three YAC
clones Y39A9, Y51D6 and Y53F1 on LGI. unc-14 had been mapped around the
region common to the three YACs. This region is also shared by several
cosmids, and one of them R05H7 was shown to complement an unc-14
mutation by Tom Barnes, according to the information in ACEDB.
Therefore, we thought that UNC-14 could possibly interact with UNC-51.
To examine this possibility, we hybridized each of several cosmids
around the unc-14 region with the cDNA clone. Only R05H7 strongly
hybridized. We confirmed that this cosmid rescued unc-14(e57)
mutation. A 17kb genomic fragment of R05H7, which hybridizes with the
cDNA clone, also perfectly rescues.
The longest cDNA clone was completely sequenced, and PCR analysis
using SL1 and SL2 primers was done. Based on these results, the mRNA of
this gene has SL1 at the 5' end, is about 2.8kb excluding poly A,
and encodes a novel protein of 665 amino acid residues. At present we
think it highly possible that our cDNAs represent the unc-14 gene.
If our cDNA is unc-14 , it leads to the possibility that UNC-14
directly interacts with UNC-51 as a substrate or a regulator. If not,
our cDNA will represent a new gene whose product can interact with
UNC-51. Interestingly, the C-terminal region of UNC-51 can interact
with UNC-51 itself. This suggests that UNC-51 acts as a dimer, or that
the C-terminal region regulates the kinase activity. Now, we plan to
examine this gene in unc-14 mutants, its expression by GFP fusion,
and direct interaction of the product with UNC-51.