Copy Link
Add to Bookmark
Report
dictyNews Volume 31 Number 12
dictyNews
Electronic Edition
Volume 31, number 12
October 18, 2008
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.
Upon publication of your paper, please send strains and plamids to
the Dicty Stock Center. For more information see
http://dictybase.org/StockCenter/Deposit.html.
Back issues of dictyNews, the Dicty Reference database and other
useful information is available at dictyBase - http://dictybase.org.
=========
Abstracts
=========
Dictyostelium discoideum Paxillin Regulates Actin-Based Processes
M. Berenice Duran, Asif Rahman, Max Colten and Derrick Brazill
Department of Biological Sciences, Center for the Study of Gene Structure
and Function
Hunter College of the City University of New York, New York, NY 10021
Protist, in press
Paxillin is a key player in integrating the actin cytoskeleton with adhesion,
and thus is essential to numerous cellular processes including proliferation,
differentiation and migration in animal cells. PaxB, the Dictyostelium
discoideum orthologue of paxillin, has been shown to be important for
adhesion and development, much like its mammalian counterpart. Here, we
use the overproduction of PaxB to gain better insight into its role in
regulating the actin cytoskeleton and adhesion. We find that PaxB
overexpressing (PaxBOE) cells can aggregate and form mounds normally, but
are blocked in subsequent development. This arrest can be rescued by
addition of wild-type cells, indicating a non-cell autonomous role for PaxB.
PaxBOE cells also have alterations in several actin-based processes,
including adhesion, endocytosis, motility and chemotaxis. PaxBOE cells
exhibit an EDTA-sensitive increase in cell-cell cohesion, suggesting that
PaxB-mediated adhesion is Ca2+ or Mg2+ dependent. Interestingly, cells
overexpressing paxB are less adhesive to the substratum. In addition,
PaxBOE cells display decreased motility under starved conditions, decreased
endocytosis, and are unable to efficiently chemotax up a folate gradient.
Taken together, the data suggest that proper expression of PaxB is vital
for the regulation of development and actin-dependent processes.
Submitted by: Derrick Brazill [Brazill@GENECTR.HUNTER.CUNY.EDU]
--------------------------------------------------------------------------------
Regulation of Rap1 Activity is Required for Differential Adhesion,
Cell Type Patterning and Morphogenesis in Dictyostelium
Katie Parkinson1, Parvin Bolourani2, David Traynor3, Nicola L. Aldren1,
Robert R. Kay3, Gerald Weeks2 and Christopher R.L. Thompson1*
1Faculty of Life Sciences, University of Manchester, Michael Smith Building,
Oxford Road, Manchester, M13 9PT
2Department of Microbiology and Immunology, University of British Columbia,
Vancouver, British Columbia, V6T 1Z3
3MRC Laboratory of Molecular Biology, Hills Road, Cambridge, CB2 2QH
*Author for correspondence (christopher.thompson@manchester.ac.uk)
Journal of Cell Science, in press
Regulated cell adhesion and motility play important roles during growth,
development and tissue homeostasis. Consequently great efforts have been
made to identify genes that control these processes. One candidate is Rap1,
as it has been implicated in the regulation of adhesion and motility in cell
culture. In order to further study the role of Rap1 during multicellular
development, we have generated a mutant in a potential Rap1 GTPase activating
protein (RapGAPB) in Dictyostelium. rapGAPB- cells have increased levels of
active Rap1 compared to wild type cells, indicating that RapGAPB regulates
Rap1 activity. Furthermore, rapGAPB- cells exhibit hallmark phenotypes of
other known mutants with hyperactivated Rap1, including increased substrate
adhesion and abnormal F-actin distribution. However, unlike these other
mutants, rapGAPB- cells do not exhibit impaired motility or chemotaxis,
indicating that RapGAPB may only regulate specific roles of Rap1.
Importantly, we also found that RapGAPB regulates Rap1 activity during
multicellular development and is required for normal morphogenesis.
Firstly, streams of aggregating rapGAPB- cells break up as a result of
decreased cell-cell adhesion. Secondly, rapGAPB- cells exhibit cell
autonomous defects in prestalk cell patterning. Using cell type specific
markers, we demonstrate that RapGAPB is required for the correct sorting
behaviour of different cell types. Finally, we show that inactivation
of RapGAPB affects prestalk and prespore cell adhesion. We therefore
propose that a possible mechanism for RapGAPB regulated cell sorting is
through differential adhesion.
Submitted by: Chris Thompson [christopher.thompson@manchester.ac.uk]
--------------------------------------------------------------------------------
Microarray phenotyping places cyclase associated protein CAP at the crossroad
of signaling pathways reorganizing the actin cytoskeleton in Dictyostelium
Hameeda Sultana(1), Girish Neelakanta(1), Ludwig Eichinger(1),
Francisco Rivero(1,2),
Angelika A. Noegel(1)
1Center for Biochemistry, Medical Faculty, Center for Molecular Medicine
Cologne (CMMC) and Cologne Excellence Cluster on Cellular Stress Responses in
Aging-Associated Diseases (CECAD), University of Cologne, 50931 Köln, Germany.
2The Hull York Medical School and Department of Biological Sciences,
University of Hull, Hull HU6 7RX, UK
Exp. Cell Res., in press
Large-scale gene expression analysis has been applied recently to uncover
groups of genes that are co-regulated in particular processes. Here we
undertake such an analysis on CAP, a protein that participates in the
regulation of the actin cytoskeleton and in cAMP signaling in Dictyostelium.
Microarray analysis revealed that loss of CAP altered the expression of many
cytoskeletal components. One of these, the Rho GDP-dissociation inhibitor
RhoGDI1, was analyzed further. RhoGDI1 null cells expressed lower amounts
of CAP, which failed to accumulate predominantly at the cell cortex. To
further position CAP in the corresponding signal transduction pathways we
studied CAP localization and cellular functioning in mutants that have
defects in several signaling components. CAP showed correct localization
and dynamics in all analyzed strains except in mutants with deficient cAMP
dependent protein kinase A activity, where CAP preferentially accumulated
in crown shaped structures. Ectopic expression of CAP improved the efficiency
of phagocytosis in Gbeta-deficient cells and restored the pinocytosis,
morphology and actin distribution defects in a PI3 kinase double mutant
(pi3k1/2 null). Our results show that CAP acts at multiple crossroads and
links signaling pathways to the actin cytoskeleton either by physical
interaction with cytoskeletal components or through regulation of their
gene expression.
Submitted by: Francisco Rivero [f.rivero-crespo@hull.ac.uk]
--------------------------------------------------------------------------------
Spatiotemporal Regulation of Ras Activity Provides Directional Sensing
Sheng Zhang1, Pascale G. Charest1, and Richard A. Firtel
1. Co-first authors
Section of Cell and Developmental Biology
Division of Biological Sciences
Center for Molecular Genetics
University of California, San Diego
9500 Gilman Drive
La Jolla, California 92093-0380
Curr. Biol., in press
Cells’ ability to detect and orient themselves in chemoattractant
gradients has been the subject of numerous studies, but
the underlying molecular mechanisms remain largely unknown
[1]. Ras activation is the earliest polarized response
to chemoattractant gradients downstream from heterotrimeric
G proteins in Dictyostelium, and inhibition of Ras signaling
results in directional migration defects [2]. Activated
Ras is enriched at the leading edge, promoting the localized
activation of key chemotactic effectors, such as PI3K and
TORC2 [2–5]. To investigate the role of Ras in directional
sensing, we studied the effect of its misregulation by using
cells with disrupted RasGAP activity. We identified an ortholog
of mammalian NF1, DdNF1, as a major regulator of Ras
activity in Dictyostelium. We show that disruption of nfaA
leads to spatially and temporally unregulated Ras activity,
causing cytokinesis and chemotaxis defects. By using unpolarized,
latrunculin-treated cells, we show that tight regulation
of Ras is important for gradient sensing. Together, our
findings suggest that Ras is part of the cell’s compass and
that the RasGAP-mediated regulation of Ras activity affects
directional sensing.
Submitted by: Rick Firtel [rafirtel@ucsd.edu]
==============================================================
[End dictyNews, volume 31, number 12]
