Copy Link
Add to Bookmark
Report
dictyNews Volume 17 Number 10
Dicty News
Electronic Edition
Volume 17, number 10
October 27, 2001
Please submit abstracts of your papers as soon as they have been
accepted for publication by sending them to dicty@northwestern.edu.
Back issues of Dicty-News, the Dicty Reference database and other useful
information is available at DictyBase--http://dictybase.org.
=============
Abstracts
=============
The NF-kB like DNA binding activity observed in Dictyostelium nuclear
extracts is due to the GBF transcription factor
Franois TRAINCARD 1, Eleonora PONTE 1, Jason PUN 2, Barrie COUKELL 2 and
Michel VERON 1
1: Unit de Rgulation enzymatique des Activits cellulaires, CNRS FRE 2364,
Institut Pasteur, 25 rue du Dr. Roux, 75724, Paris Cedex 15, France.
2: York University, 4700 Keele St., Toronto, Ontario, M3J 1P3, Canada.
Accepted in Journal of Cell Science
We have previously reported that a NF-kB transduction pathway was likely
to be present in the cellular slime mold Dictyostelium discoideum
(Traincard, F., Ponte, E., Pun, J., Coukell, B. and Vron, M., 1999,
J. Cell Science, 112, 3529-3535). This conclusion was based on several
observations, including the detection of developmentally regulated DNA
binding proteins in Dictyostelium nuclear extracts, which bound to bona
fide kB sequences. We have now performed additional experiments that
demonstrate that the protein responsible for this NF-kB like DNA binding
activity is the Dictyostelium GBF (G box regulatory element Binding Factor)
transcription factor. This result, along with the fact that no sequence with
significant similarity to components of the mammalian NF-kB pathway can be
found in Dictyostelium genome, now almost entirely sequenced, led us to
reconsider our previous conclusion on the occurrence of a NF-kB signal
transduction pathway in Dictyostelium.
-----------------------------------------------------------------------------
GTP[gamma]S regulation of a twelve transmembrane guanylyl
cyclase is retained after mutation to an adenylyl cyclase
Jeroen Roelofs, Harrit M. Loovers and Peter J.M. Van
Haastert
Department of Biochemistry, University of Groningen,
Nijenborgh 4,9747AG Groningen, the Netherlands
J. Biol. Chem., in press
ABSTRACT
DdGCA is a Dictyostelium guanylyl cyclase with a topology
typical for mammalian adenylyl cyclases containing twelve
transmembrane spanning regions and two cyclase domain. In
Dictyostelium cells heterotrimeric G-proteins are essential for
guanylyl cyclase activation by extra-cellular cAMP. In lysates,
guanylyl cyclase activity is strongly stimulated by GTPgS, which
is also a substrate of the enzyme. DdGCA was converted to an
adenylyl cyclase by introducing three point mutations.
Expression of the obtained DdGCAkqd in adenylyl cyclase
defective cells restored the phenotype of the mutant. GTPgS
stimulated the adenylyl cyclase activity of DdGCAkqd with similar
properties as the wildtype enzyme (decrease of Km and increase
of Vmax). This demonstrates that GTPgS-stimulation is
independent of substrate specificity. Furthermore, GTPgS- activation
of DdGCAkqd is retained in several null mutants of Ga
and Gb proteins, indicating that GTPgS-activation is not
mediated by a heterotrimeric G-protein, but possibly by a
monomeric G-protein.
-----------------------------------------------------------------------------
Single molecule analysis of chemotactic signaling in Dictyostelium cells
Masahiro Ueda, Yasushi Sako, Toshiki Tanaka, Peter Devreotes & Toshio
Yanagida
Science 294, 864-867.
Single-molecule imaging techniques were used to reveal the binding of
individual cyclic adenosine 3',5'-monophosphate molecules to heterotrimeric
guanine nucleotide-binding protein coupled receptors on the surface of
living Dictyostelium cells. The binding sites were uniformly distributed
and diffused rapidly in the plane of the membrane. The probabilities of
individual association and dissociation events were greater for receptors
at the anterior end of the cell. Agonist-induced receptor phosphorylation
had little effect on any of the monitored properties, whereas G-protein
coupling influenced the binding kinetics. These observations illustrate
the dynamic properties of receptors involved in gradient sensing and
suggest that these may be polarized in chemotactic cells.
-----------------------------------------------------------------------------
Calreticulin and Calnexin in the Endoplasmic Reticulum Are Important for
Phagocytosis
Annette Mueller-Taubenberger*, Andrei N. Lupas, Hewang Li*, Mary Ecke*,
Evelyn Simmeth*, and Guenther Gerisch*
* Max-Planck-Institut fuer Biochemie, Martinsried, Germany
SmithKline Beecham Pharmaceuticals UP1345, Collegeville, PA, USA
EMBO Journal, in press.
Abstract
Calreticulin and calnexin are Ca2+-binding proteins with chaperone activity
in the endoplasmic reticulum. These proteins have been eliminated by gene
replacement in Dictyostelium, the only microorganism known to harbor both
proteins; family members in Dictyostelium are located at the base of
phylogenetic trees. In double mutants lacking calreticulin and calnexin
a dramatic decline in the rate of phagocytosis was observed, whereas only
mild changes occurred in single mutants. Dictyostelium cells are
professional phagocytes, capable of internalizing particles by a sequence
of activities: adhesion of the particle to the cell surface, actin-
dependent outgrowth of a phagocytic cup, and separation of the phagosome
from the plasma membrane. In the double-null mutants, particles still
adhered to the cell surface, but the outgrowth of phagocytic cups was
compromised. GFP-tagged calreticulin and calnexin, expressed in wild type
cells, revealed a direct link of the ER to the phagocytic cup enclosing a
particle, such that the Ca2+-storage capacity of calreticulin and calnexin
might directly modulate activities of the actin system during particle
uptake.
-----------------------------------------------------------------------------
[End Dicty News, volume 17, number 10]
