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dictyNews Volume 26 Number 10
dictyNews
Electronic Edition
Volume 26, number 10
April 7, 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 of the major spliceosomal RNAs in Dictyostelium discoideum
reveals developmentally regulated U2 variants and polyadenylated snRNAs.
Andrea Hinas(1), Pontus Larsson(2), Lotta Avesson(1), Leif A. Kirsebom(2),
Anders Virtanen(2), and Fredrik Soederbom(1)
(1)Department of Molecular Biology, Biomedical Center, Swedish University
of Agricultural Sciences, Box 590, SE-75124 Uppsala, Sweden
(2)Department of Cell and Molecular Biology, Biomedical Center, Uppsala
University, Box 596, SE-75124 Uppsala, Sweden.
Eukaryotic Cell, in press
Most eukaryotic mRNAs depend upon precise removal of introns by the
spliceosome, a complex of RNAs and proteins. Splicing of pre-mRNA is known
to take place in Dictyostelium discoideum and we previously isolated the U2
spliceosomal RNA experimentally. In this study, we identified the remaining
major spliceosomal RNAs in Dictyostelium by a bioinformatical approach.
Expression was verified from 17 genes encoding these small nuclear RNAs
(snRNAs). All these genes are preceded by a putative non-coding RNA gene
promoter. Immunoprecipitation showed that snRNAs U1, U2, U4, and U5, but not
U6, carry the conserved trimethylated 5Õ cap structure. A number of
divergent U2 species are expressed in Dictyostelium. These RNAs carry the
U2 RNA hallmark sequence and structure motifs but have an additional
predicted stem-loop structure at the 5Õ end. Surprisingly, and in contrast
to the other spliceosomal RNAs in this study, the new U2 variants were
enriched in the cytoplasm and were developmentally regulated. Furthermore,
all of the snRNAs could also be detected as polyadenylated species and
polyadenylated U1 RNA was demonstrated to be located in the cytoplasm.
Submitted by: Fredrik Soederbom [fredde@xray.bmc.uu.se]
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Aberrant stalk development and breakdown of tip dominance in
Dictyostelium cell lines with RNAi-silenced expression of calcineurin B
Katrina Boeckeler1,2, Gilbert Tischendorf1, Rupert Mutzel1 and
Barbara Weissenmayer1
1Institut fr Biologie Ð Mikrobiologie, Fachbereich Biologie, Chemie,
Pharmazie, Freie Universitaet Berlin, Koenigin-Luise-Strasse 12-16,
14195 Berlin, Germany and 2University College London, Department of Biology,
Gower Street, London, Wc1 E6BT, UK
BMC Developmental Biology, in press
Background: Calcineurin, the Ca2+/calmodulin-dependent protein phosphatase,
plays important roles in various cellular processes in lower and higher
eukaryotes. Here we analyze the role of calcineurin in the development of
Dictyostelium discoideum by RNAi-mediated manipulation of its
expression.
Results: The cnbA gene of Dictyostelium discoideum which encodes the
regulatory B subunit (CNB) of calcineurin was silenced by RNAi. We found a
variety of silencing levels of CNB in different recombinant cell lines.
Reduction of CNB expression in a given cell line was correlated with
developmental aberrations. Cell lines with strongly reduced protein levels
developed slower than wild type cells and formed short stalks and spore
heads with additional tips. Formation of short stalks results from
incomplete vacuolization of prestalk cells during terminal differentiation.
Expression of the stalk-specific gene ecmB was reduced in mutant cells.
Aberrant stalk development is a cell autonomous defect, whereas the
breakdown of tip dominance can be prevented by the presence of as low as
10% wild type cells in chimeras.
Conclusion: Silencing of calcineurin B in Dictyostelium by expression of
RNAi reveals an unexpected link between increased intracellular calcium
levels, possibly triggered by the morphogen DIF, activation of calcineurin,
and the terminal stage of morphogenesis.
Submitted by: Barbara Weissenmayer [bweiss@zedat.fu-berlin.de]
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Transcriptional pulsing of a developmental gene
Jonathan R. Chubb, Tatjana Trcek, Shailesh M. Shenoy and Robert H. Singer
Division of Cell and Developmental Biology, School of Life Sciences,
University of Dundee, Dundee, DD1 5EH, UK and Department of Anatomy and
Structural Biology, Albert Einstein College of Medicine, The Bronx,
New York 10461, USA.
Current Biology, in press
It has not been possible to view the transcriptional activity of a single
gene within a living eukaryotic cell. It is therefore unclear how long and
how frequently a gene is actively transcribed, how this is modulated during
differentiation and how transcriptional events are dynamically coordinated
in cell populations. Using an in vivo RNA detection technique, we have
directly visualised transcription of an endogenous developmental gene. We
found discrete 'pulses' of gene activity, that turn on and off at irregular
intervals. Surprisingly, the length and height of these pulses were
consistent throughout development. However, there was strong developmental
variation in the proportion of cells recruited to the expressing pool.
Cells were more likely to re-express than to initiate new expression,
indicating we directly observe a transcriptional memory. In addition, we
used a clustering algorithm to reveal synchronous transcription initiation
in neighbouring cells. This study represents the first direct visualisation
of transcriptional pulsing in eukaryotes. Discontinuity of transcription may
allow greater flexibility in the gene expression decisions of a cell.
Submitted by: Jonathan Chubb [j.chubb@dundee.ac.uk]
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[End dictyNews, volume 26, number 10] 
