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dictyNews Volume 23 Number 04
Dicty News
Electronic Edition
Volume 23, number 4
July 30, 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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Dictyostelium calcium-binding protein 4a interacts with NumA, a BRCT-domain
protein that regulates nuclear number
Michael A. Myre & Danton H. OâDay*
Department of Biology, University of Toronto at Mississauga, Mississauga,
ON. Canada
Biochemical and Biophysical Research Communications, in press
Nucleomorphin from Dictyostelium discoideum is a nuclear calmodulin binding
protein that is a member of the BRCT-domain containing cell cycle
checkpoint proteins. Two differentially expressed isoforms, NumA and NumB,
share an extensive acidic domain (DEED) that when deleted produces highly
multinucleated cells. We performed a yeast two-hybrid screen of a
Dictyostelium cDNA library using NumA as bait. Here we show that
nucleomorphin interacts with calcium binding protein 4a (CBP4a) in a
Ca2+-dependent manner. Further deletion analysis suggests this interaction
requires residues found within the DEED domain. NumA and CBP4a mRNAs are
expressed at the same stages of development. CBP4a belongs to a large family
of Dictyostelium CBPs, for which no cellular or developmental functions had
previously been determined. Since the interaction of CBP4a with NumA
requires the DEED domain, this suggests that CBP4a may respond to
Ca2+-signalling through modulating factors that might function in concert
to regulate nuclear number.
Submitted by: M. Myre" [mmyre@utm.utoronto.ca]
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Rac regulation of chemotaxis and morphogenesis in Dictyostelium
Kyung Chan Park1, Francisco Rivero2, Ruedi Meili1, Susan Lee1, Fabio Apone1,3,
and Richard A. Firtel1,4
1Section of Cell and Developmental Biology, Division of Biological Sciences
Center for Molecular Genetics, University of California, San Diego
9500 Gilman Drive, La Jolla, CA 92093-0634
2Zentrum fr Biochemie der Medizinischen Fakultt, Universitt zu Kln
Joseph-Stelzmann-Strasse 52
50931 Kln, Germany
EMBO J, in press
Chemotaxis requires localized F-actin polymerization at the site of the
plasma membrane closest to the chemoattractant source, a process controlled
by Rac/Cdc42 GTPases. We identify Dictyostelium RacB as an essential mediator
of this process. RacB is activated upon chemoattractant stimulation,
exhibiting biphasic kinetics paralleling F-actin polymerization. racB null
cells have strong chemotaxis and morphogenesis defects and a severely reduced
chemoattractant-mediated F-actin polymerization and PAKc activation. RacB
activation is partly controlled by the PI3K pathway. pi3k1/2 null cells and
wild-type cells treated with LY294002 exhibit a significant reduced second
peak of RacB activation and that is linked to pseudopod extension, whereas
a PTEN hypomorph exhibits elevated RacB activation. We identify a RacGEF,
RacGEF1, which has a specificity for RacB in vitro. racgef1 null cells
exhibit reduced RacB activation and cells expressing mutant RacGEF1 proteins
display chemotaxis and morphogenesis defects. RacGEF1 localizes to sites of
F-actin polymerization. Inhibition of this localization reduces RacB
activation, suggesting a feedback loop from RacB via F-actin polymerization
to RacGEF1. Our findings provide a critical linkage between chemoattractant
stimulation, F-actin polymerization, and chemotaxis in Dictyostelium.
Submitted by: Rick Firtel [rafirtel@ucsd.edu]
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A cell number counting factor regulates Akt/ PKB to regulate group size in
Dictyostelium
Tong Gao1, David Knecht2, Lei Tang3, R. Diane Hatton1,
and Richard H. Gomer1,3
1HHMI/ 3Rice University, Houston, TX 77005-1892 and 2University of
Connecticut, Storrs, Connecticut 06269-3125
Eukaryotic Cell, in press
Little is known about how individual cells can organize themselves to form
structures of a given size. During development, Dictyostelium discoideum
aggregates in dendritic streams and forms groups of ~20,000 cells.
D. discoideum regulates group size by secreting and simultaneously sensing
a multi-protein complex called counting factor (CF). If there are too
many cells in a stream, the associated high concentration of CF will
decrease cell-cell adhesion and increase cell motility, causing aggregation
streams to break up. The pulses of cAMP that mediate aggregation cause a
transient translocation of the Akt/PKB kinase to the leading edge of the
plasma membrane and a concomitant activation of the kinase activity, which
in turn stimulates motility. We found that countinø cells (which lack
bioactive CF) and wild-type cells starved in the presence of anti-countin
antibodies (which block CF activity) showed a decreased level of
cAMP-stimulated Akt/PKB membrane translocation and kinase activity compared
to parental wild type cells. Recombinant countin has the bioactivity of
CF, and a 1-minute treatment of cells with recombinant countin potentiated
Akt/PKB translocation to membranes and Akt/PKB activity. Western blots
of total cell lysates indicated that countin does not affect the total
level of Akt/PKB. Fluorescence microscopy of cells expressing an Akt/PKB
PH-GFP fusion protein indicated that recombinant countin and anti-countin
antibodies do not obviously alter the distribution of Akt/PKB PH-GFP when
it translocates to the membrane. Our data indicate that CF increases
motility by potentiating the cAMP-stimulated activation and translocation
of Akt/PKB.
Submitted by: Richard Gomer [richard@rice.edu]
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[End Dicty News, volume 23, number 4]
