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dictyNews Volume 26 Number 03
dictyNews
Electronic Edition
Volume 26, number 3
January 20, 2006
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 dictyNews, the Dicty Reference database and other
useful information is available at dictyBase - http://dictybase.org.
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Abstracts
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Identification and characterization of an 8 kDa light chain associated with
Dictyostelium discoideum MyoB, a class I myosin
Scott W. Crawley, Marc A. de la Roche, Sheu-Fen Lee, Zhihao Li, Seth Chityat,
Steven P. Smith, and Graham P. Cote
Biochemistry Dept., Queen's University, Kingston, Ontario K7L 3N6
Journal of Biological Chemistry, in press
Dictyostelium discoideum MyoB is a single-headed class I myosin. Analysis of
purified MyoB by SDS-PAGE indicated the presence of an ~9 kDa light chain. A
tryptic digest of MyoB yielded a partial sequence for the light chain that
exactly matched a sequence in a 73-amino acid, 8,296-Da protein (dictyBase
DDB0188713). This protein, termed MlcB, contains two EF-hand motifs and
shares ~30% sequence identity with the N- and C-terminal lobes of calmodulin.
Flag-MlcB expressed in Dictyostelium co-immunoprecipitated with MyoB, but
not with the related class I myosins MyoC and MyoD. Recombinant MlcB bound
Ca2+ with a Kd value of 0.2 µM and underwent a Ca2+-induced change in
conformation that increased alpha-helical content and surface
hydrophobicity. Mutational analysis showed that the first EF-hand was
responsible for Ca2+ binding. In the presence and absence of Ca2+ MlcB was
a monomer in solution and bound to a MyoB IQ motif peptide with a Kd value
of ~0.5 µM. A MyoB head-neck construct with a Ser to Glu mutation at the
TEDS site bound MlcB and displayed an actin-activated MgATPase activity
that was insensitive to Ca2+. We conclude that MlcB represents a novel type
of small myosin light chain that binds to IQ motifs in a manner comparable
to a single lobe of a typical four EF-hand protein.
Submitted by: Graham Cote [coteg@post.queensu.ca]
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The C-module DNA-binding factor mediates expression of the Dictyostelium
aggregation-specific adenylyl cyclase ACA
Oliver Siol (1) Theodor Dingermann (1) and Thomas Winckler (1,2)
(1) Institut fuer Pharmazeutische Biologie, Universitaet Frankfurt/M.
(Biozentrum), Frankfurt am Main, Germany, and (2) Lehrstuhl fuer
Pharmazeutische Biologie, Universitaet Jena, Germany
Eukaryot. Cell, in press
Aggregation of Dictyostelium discoideum amoebae into multicellular
structures is organized by cyclic AMP, which acts both as a chemoattractant,
second messenger, and morphogen. Aggregation of D. discoideum cells depends
on the expression of adenylyl cyclase ACA, which provides extracellular cAMP
for signal relay and intracellular cAMP for the induction of genes required
at multicellular stages. We have identified a DNA-binding activity specific
for a highly A+T-enriched motif in the upstream region of the ACA-encoding
gene, acaA. The factor shows DNA-binding characteristics very similar to
that of C-module-binding factor (CbfA). Although CbfA was originally
identified as a putative regulator of the activity of D. discoideum
retrotransposon TRE5-A, it was also found to be essential for aggregation
of D. discoideum cells. The identified DNA-binding activity was absent in
mutant cells depleted for CbfA, and CbfA could be precipitated using an acaA
promoter fragment. We propose that CbfA binds to the acaA promoter to
provide a basal transcription activity that is required for induction of
ACA expression after the onset of D. discoideum development.
Submitted by: Thomas Winckler [t.winckler@uni-jena.de]
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[End dictyNews, volume 26, number 3] 
