Worm Breeder's Gazette 14(1): 38 (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 Neurology, Baylor College of Medicine, Houston, TX.
We are interested in determining the possible use of C. elegans
as an animal model to study the protein(s) indicated in the human
disease Myotonic Dystrophy, (DM). We therefore performed a computer
search of the dbEST database to locate cDNAs that were similar in
sequence to the DM protein kinase gene, (DMPK). We found four
different sequences with protein sequence similarities ranging from 50
to 95% over the short regions that had been sequenced. One cDNA
(Accession # T01329) in the databases was originally found by Anthony
Kerlavage at the Institute for Genomic Research in Gaithersburg, MD.
The clone, CEESP52, came from a C. elegans embryonic cDNA library. One
open-reading frame of the clone contained a protein sequence that
matches at a score of 71% identity and 95% similarity to part of the
catalytic domain of the human DM kinase gene. We have hybridized a
probe made from CEESP52 to the Yac contig library and determined that
the gene containing the coding sequences is found on Yac Y38B5. Yac
Y38B5 is covered by known cosmids except for an approximately 20kb gap
in the cosmid contigs. A Southern indicated that the region
sequenced did not hybridize to any of the cosmids, therefore we believe
that at least part, if not all, of the gene is located in the gap
region. We have run pulse field gels and isolated Yac Y38B5 DNA and
will be screening subclones of the Yac for the corresponding gene.
We will then perform rescue experiments of known mutations or construct
our own mutant.
As another approach to locating DMPK homologs we have used is
employing a monoclonal and polyclonal antibodies made to the human DMPK
protein on Western blots containing C. elegans homogenates. The
monoclonal antibody identified an approximately 80kDa band on the
Western, whereas the polyclonal detected 5 different bands. We are in
the process of isolating these genes from a lgt11 embryonic library.
Below is a translated protein sequence match of the 4 closest
homologs to DMPK found in the dbEST database over a overlapping 100
amino acid region. Two other cDNA sequences in the dbEST database have
good similarity scores to DMPK equivalent to the C. elegans Ndr peptide
score, but are not in the same region. Both have been mapped and are
found on chromosome I (1,2). The partial sequence of the Ndr protein
has been recently published and has homologs in both Drosophila and
humans (3).
Upper Sequence : Human DMPK (all matches to the Human DMPK)
2nd Sequence : Accession # T01329 peptide
Match Percentage : 71%
3rd Sequence : C. elegans Ndr peptide
Match Percentage : 45%
4th Sequence : Accession # D37290 peptide
Match Percentage : 35%
. : . : . : . : . :
1 DFEILKVIGRGAFSEVAVVKMKQTGQVYAMKIMNKWDMLKRGEVSCFREE
|||:|||||:|||:|||||:|: |::|||||:|||:|:||:| :|||||
1 DFEVLKVIGKGAFGEVAVVRMRGVGEIYAMKILNKWEMVKRAETACFREE
:| |||||||||:|| :|: :||::|||||: | :|: : : : | |
1 HFKSLKVIGRGAFGEVRLVQKHDTGHIYAMKILRKSEMVEKEQTAHVRAE
:| :|:|:|:||: :| :|: | :||||:: | :| : : |
1 NFALLRVLGKGAYGKVFLVRKDHNTIYAMKVLRKTRVLTKQKXLEHTMAE
. : . : . : . : .
51 RDVLVNGDRRWITQLHFAFQDENYLYLVMEYYVGGDLLTLLSKFGERIP
||||| |||||||:||:|||||: ||:||:||:|||:||||||| ::||
51 RDVLVYGDRRWITNLHYAFQDEKNLYFVMDYYIGGDMLTLLSKFVDHIP
||:| ::| |: :: ::||| : ||||||: |||::||| | :
51 RDILSEADCDWVVKMYYSFQDYSNLYLVMEFLPGGDMMTLLIKKATLTE
| || :: | :||| : |::|||| ||:| | | |
51 RQVLERLRTPFLVNLXYAFQTDXKLHIVMEYVRGGELFTHLCSRGH
1) Gilchrist, E. and Baillie, D., (1995) A C. elegans
Myotonic Dystrophy gene homologue. Abstracts of the 10th International
C. Elegans meeting. University of Wisconsin, Madison. P233.
2) Wissmann, A., McGhee, J., Mains, P., (1995) Let-502 is a
homolog of human Myotonic Dystrophy kinase. Abstractions of the 10th
International C. elegans meeting. University of Wisconsin, Madison.
P548.
3) Millward, T., Cron, P., Hemmings, B., (1995) Molecular
cloning and characterization of a conserved nuclear serine(threonine)
protein kinase. Proc. Natl .Acad. Sci. USA 92:5022-6.