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dictyNews Volume 31 Number 17
dictyNews
Electronic Edition
Volume 31, number 17
December 5, 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.
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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Subcellular localization of ammonium transporters in
Dictyostelium discoideum
Janet H. Kirsten (1), Yanhua Xiong (1), Carter T. Davis (2), Charles
K. Singleton (1)
(1) Department of Biological Sciences, Vanderbilt University, VU
Station B 351634, Nashville TN 37235-1634, USA
(2) LSU School of Medicine – New Orleans, 2020 Gravier Street, New
Orleans, LA 70112, USA
BMC Cell Biology, in press
Background
With the exception of vertebrates, most organisms have plasma membrane
bound ammonium transporters which primarily serve to import a source
of nitrogen for nutritional purposes. Dictyostelium discoideum has three
ammonium transporters, Amts A, B and C. Our present work used
fluorescent fusion proteins to determine the cellular localization of the
Amts and tested the hypothesis that the transporters mediate removal of
ammonia generated endogenously from the elevated catabolic activity
common to many protists.
Results
Using RFP and YFP fusion constructs driven by the actin 15 promoter,
we found that the three ammonium transporters were localized on the
plasma membrane and on subcellular organelles. AmtA and AmtB were
localized on the membranes of endolysosomes and phagosomes, with
AmtB further localized on the membranes of contractile vacuoles. AmtC
alsowas localized on subcellular organelles when it was stabilized by
coexpression with either the AmtA or AmtB fusion transporter. The
three ammonium transporters exported ammonia linearly with regard to
time during the first 18 hours of the developmental program as
revealed by reduced export in the null strains. The fluorescently
tagged transporters rescued export when expressed in the null strains,
and thus they were functional transporters.
Conclusions
Unlike ammonium transporters in most organisms, which import NH3/NH4+
as a nitrogen source, those of Dictyostelium export ammonia/ammonium
as a waste product from extensive catabolism of exogenously derived
and endogenous proteins. Localization on proteolytic organelles and on
the neutral contractile vacuole suggests that Dictyostelium ammonium
transporters may have unique subcellular functions and play a role in
the maintenance of intracellular ammonium distribution. A lack of
correlation between the null strain phenotypes and ammonia excretion
properties of the ammonium transporters suggests that it is not the
excretion function that is important for coupling ammonia levels to
the slug versus culmination choice, but rather a sensor and/or
signaling function of these proteins that is important.
Submitted by: Janet Kirsten [Jan.Kirsten@vanderbilt.edu]
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Role of an expansin-like molecule in Dictyostelium morphogenesis and
regulation of its gene expression by the STAT protein Dd-STATa
Shun Ogasawara1, Nao Shimada1, 2 and Takefumi Kawata1, 3
1 Department of Biology, Faculty of Science, Toho University, 2-2-1 Miyama,
Funabashi, Chiba 274-8510, Japan
2Present Address: Department of Basic Science, Graduate School of Arts and
Sciences, University of Tokyo, 3-8-1 Komaba, Meguro-ku, Tokyo, 153-8902, Japan
3Corresponding author
E-mail: tkawata@bio.sci.toho-u.ac.jp
Devleop. Growrh Differ., in press
Expansins are proteins involved in plant morphogenesis, exerting their
effects on cellulose to extend cell walls. Dictyostelium is an organism
that possesses expansin-like molecules, but their functions are not
known. In this study, we analysed the expL7 (expansin-like 7) gene,
which has been identified as a putative target of Dd-STATa, a
Dictyostelium homologue of the metazoan signal transducer and
activator of transcription (STAT) proteins. Promoter fragments of the
expL7 were fused to a lacZ reporter and the expression patterns
determined. As expected from the behaviour of the endogenous
expL7 gene, the expL7/lacZ fusion gene was downregulated in
Dd-STATa null slugs. In the parental strain, the expL7 promoter was
activated in the anterior tip region. Mutational analysis of the promoter
identified a sequence that was necessary for expression in tip
cells. In addition, an activator sequence for pstAB cells was
identified. These sequences act in combination with the repressor
region to prevent ectopic expL7 expression in the prespore and
prestalk regions of the slug and culminant. Although the expL7 null
mutant showed no phenotypic change, the expL7 overexpressor
showed aberrant stalk formation. These results indicate that the
expansin-like molecule is important for morphogenesis in Dictyostelium.
Submitted by: Takefumi Kawata [tkawata@bio.sci.toho-u.ac.jp]
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[End dictyNews, volume 31, number 17]
