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dictyNews Volume 22 Number 16
Dicty News
Electronic Edition
Volume 22, number 16
June 18, 2004
Please submit abstracts of your papers as soon as they have been
accepted for publication by sending them to dicty@northwestern.edu
or by using the form at
http://dictybase.org/db/cgi-bin/dictyBase/abstract_submit.
Back issues of Dicty-News, the Dicty Reference database and other
useful information is available at dictyBase - http://dictybase.org.
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Abstracts
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Novel development rescuing factors (DRFs) secreted by the developing
Dictyostelium cells, that are involved in the restoration of a mutant
lacking MAP-kinase ERK2
Masatsune Tsujioka1,2, Tatsuyoshi Yamamoto1, Christopher R. Thompson2,
Robert R. Kay2 and Mineko Maeda1*
1Department of Biology, Osaka University, Machikaneyama-cho 1-16,
Toyonaka, Osaka 560-0043, Japan and 2MRC Laboratory of Molecular
Biology, Cambridge CB2 2QH, UK
Zoological Science, in press
We found novel development rescuing factors (DRFs) secreted from developing
Dictyostelium cells, by using a mutant (erkB-) which is missing MAP-kinase
ERK2 as a test strain for bioassay. The mutant erkB- fails to undergo
multicellular morphogenesis due to impaired cAMP signaling. However, such
developmental defect can be restored by the presence of low-molecular
weight DRFs that are secreted from developing wild-type cells. We
previously showed that DIF-1 (Differentiation-Inducing Factor 1 for stalk
cells) possesses this activity, indicating a newly discovered role of DIF-1.
Surprisingly, however, the mutant dmtA-, which is incapable of DIF-1
synthesis still exerts a strong inducing activity of the multicellular
morphogenesis of erkB-. After analysis of HPLC fractions of conditioned
media prepared from both wild type Ax2 and dmtA- strains revealed that both
strains secrete at least two novel DRF activities with DIF-like mobility.
However, these activities were not derived from other DIFs such as DIF-2
and DIF-3. Identification of these DRFs found in this study would provide
insight into the mechanism by which the development of the erkB- mutant is
restored and how these factors act in the normal development of
Dictyostelium.
Submitted by: Mineko Maeda [mmaeda@bio.sci.osaka-u.ac.jp]
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Identification of Phospholipase B from Dictyostelium discoideum reveals a
new Lipase Family Present In Mammals, Flies and Nematodes, but not Yeast
Clive P. Morgan, Robert Insall*, Lee Haynes and Shamshad Cockcroft
Biochem J., in press
The social amoeba Dictyostelium discoideum exhibits high activities of
phospholipase and lysophospholipase (Ferber et al Eur.J.Biochem. 1970 14
253-257). We assayed Dictyostelium lysates to demonstrate the presence of a
highly active phospholipase B enzyme that removed both fatty-acidchains from
phosphatidylcholine (PC) and produced the water-soluble
glycerophosphorylcholine. We purified the PLB activity from Dictyostelium
cytosol using standard agarose media (size exclusion and ion-exchange), and
combined this with an affinity step using myristoylated ARF1, a protein
which has a single fatty acid at its N-terminus. Two proteins co-purified
(48kDa and 65kDa), and the 48kDa protein was digested with trypsin, peptide
fragments separated by reverse phase chromatography, and the resultant
peptides were sequenced by Edman degradation. From the peptide sequences
obtained, database searches revealed a gene, which encodes a protein of
65 kDa with unknown function. The 48 kDa, therefore appears to be a
fragment of the full length 65 kDa product. Expression of the gene in
E.coli confirmed that it encodes a PLB. Characterisation of its substrate
specificity indicated that in addition to PC deacylation, the enzyme also
hydrolysed phosphatidylinositol and phosphatidylethanolamine. The PLB
identified here is not related to existing PLBs, identified in bacteria,
fungi or mammals. There are however, genes homologous to Dictyostelium PLB
in mammals, flies, worms and Giardia but not in yeast. We therefore have
identified a novel family of intracellular PLBs.
Submitted by: Robert Insall [r.h.insall@bham.ac.uk]
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[End Dicty News, volume 22, number 16]
