dictyNews
Electronic Edition
Volume 42, number 29
December 9, 2016
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Abstracts
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Actin Binding Domain of Filamin Distinguishes Posterior from Anterior Actin
Filaments in Migrating Dictyostelium Cells
Keitaro Shibata, Akira Nagasaki, Hiroyuki Adachi, Taro Q.P. Uyeda
Biophysics and Physicobiology, in press
Actin filaments in different parts of a cell interact with specific actin binding
proteins (ABPs) and perform different functions in a spatially regulated manner.
However, the mechanisms of those spatially-defined interactions have not been
fully elucidated. If the structures of actin filaments differ in different parts of a cell,
as suggested by previous in vitro structural studies, ABPs may distinguish these
structural differences and interact with specific actin filaments in the cell. To
test this hypothesis, we followed the translocation of the actin binding domain of
filamin (ABDFLN) fused with photoswitchable fluorescent protein (mKikGR) in
polarized Dictyostelium cells. When ABDFLN-mKikGR was photoswitched in the
middle of a polarized cell, photoswitched ABDFLN-mKikGR rapidly translocated
to the rear of the cell, even though actin filaments were abundant in the front. The
speed of translocation (>3 µm/s) was much faster than that of the retrograde flow
of cortical actin filaments. Rapid translocation of ABDFLN-mKikGR to the rear
occurred normally in cells lacking GAPA, the only protein, other than actin, known
to bind ABDFLN. We suggest that ABDFLN recognizes a certain feature of actin
filaments in the rear of the cell and selectively binds to them, contributing to the
posterior localization of filamin.
submitted by: Taro Uyeda [[log in to unmask]]
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A novel HECT Ubiquitin Ligase Regulating Chemotaxis and Development in
Dictyostelium discoideum
Barbara Pergolizzi, Enrico Bracco and Salvatore Bozzaro
J Cell Sci, in press
Cyclic AMP binding to G protein-coupled receptors orchestrates chemotaxis and
development in Dictyostelium. By activating the RasC-TORC2-AKT/PKB module,
cAMP regulates cell polarization during chemotaxis. TORC2 also mediates GPCR-
dependent stimulation of adenylyl cyclase A (ACA), enhancing cAMP relay and
developmental gene expression. Thus, mutants defective in the TORC2 Pia/Rictor
subunit are impaired in chemotaxis and development. Near-saturation mutagenesis
of a Pia/Rictor mutant by random gene disruption led to selection of two suppressor
mutants, in which spontaneous chemotaxis and development were restored. PKB
phosphorylation and chemotactic cell polarization were rescued, whereas Pia/Rictor-
dependent ACA stimulation was not restored but bypassed, leading to cAMP-
dependent developmental gene expression. Knocking out the gene encoding the
adenylylcyclase B (ACB) in the parental strain showed ACB to be essential for this
process. The gene tagged in the suppressor mutants encodes a novel HECT
ubiquitin ligase, homologous to mammalian HERC1, but harbouring a pleckstrin
homology domain. Expression of the isolated HECTwt, but not HECTC5185S,
domain was sufficient to reconstitute the parental phenotype. The novel ubiquitin
ligase appears to regulate cell sensitivity to cAMP signalling and TORC2-dependent
PKB phosphorylation.
submitted by: Salvo Bozzaro [[log in to unmask]]
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Actin-Interacting Protein 1 Contributes to Intranuclear Rod Assembly in
Dictyostelium discoideum
Hellen C. Ishikawa-Ankerhold, Wioleta Daszkiewicz, Michael Schleicher, and
Annette Müller-Taubenberger
Scientific Reports, in press
Intranuclear rods are aggregates consisting of actin and cofilin that are formed
in the nucleus in consequence of chemical or mechanical stress conditions. The
formation of rods is implicated in a variety of pathological conditions, such as
certain myopathies and some neurological disorders. It is still not well understood
what exactly triggers the formation of intranuclear rods, whether other proteins are
involved, and what the underlying mechanisms of rod assembly or disassembly are.
In this study, Dictyostelium discoideum was used to examine appearance, stages of
assembly, composition, stability, and dismantling of rods. Our data show that
intranuclear rods, in addition to actin and cofilin, are composed of a distinct set
of other proteins comprising actin-interacting protein 1 (Aip1), coronin (CorA),
filactin (Fia), and the 34 kDa actin-bundling protein B (AbpB). A finely tuned
spatio-temporal pattern of protein recruitment was found during formation of rods.
Aip1 is important for the final state of rod compaction indicating that Aip1 plays
a major role in shaping the intranuclear rods. In the absence of both Aip1 and
CorA, rods are not formed in the nucleus, suggesting that a sufficient supply of
monomeric actin is a prerequisite for rod formation.
submitted by: Annette Müller-Taubenberger [[log in to unmask]]
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The long non-coding RNA transcriptome of Dictyostelium discoideum development
Rafael D. Rosengarten, Balaji Santhanam, Janez Kokosar and Gad Shaulsky
Baylor College of Medicine, Houston, TX, USA
Accepted for publication in G3: Genes, Genomes, Genetics
Dictyostelium discoideum live in the soil as single cells, engulfing bacteria and
growing vegetatively. Upon starvation, tens of thousands of amoebae enter a
developmental program that includes aggregation, multicellular differentiation,
and sporulation. Major shifts across the protein-coding transcriptome accompany
these developmental changes. However, no study has presented a global survey
of long non-coding RNAs in D. discoideum. To characterize the antisense and
long intergenic non-coding RNA transcriptome, we analyzed previously published
developmental time course samples using an RNA-sequencing library preparation
method that selectively depletes ribosomal RNAs. We detected the accumulation
of transcripts for 9,833 protein-coding messenger RNAs, 621 long intergenic non-
coding RNAs and 162 putative antisense RNAs. The non-coding RNAs were
interspersed throughout the genome, and were distinct in expression level, length
and nucleotide composition. The non-coding transcriptome displayed a temporal
profile similar to the coding transcriptome, with stages of gradual change
interspersed with larger leaps. The transcription profiles of some non-coding
RNAs were strongly correlated with known differentially expressed coding RNAs,
hinting at a functional role for these molecules during development. Examining
the mitochondrial transcriptome, we modeled two novel antisense transcripts.
We applied yet another ribosomal depletion method to a subset of the samples
to better retain tRNA transcripts. We observed polymorphisms in tRNA anticodons
that suggested a post-transcriptional means by which D. discoideum compensates
for codons missing in the genomic complement of tRNAs. We concluded that the
prevalence and characteristics of long non-coding RNAs indicate these molecules
are relevant to the progression of molecular and cellular phenotypes during
development.
submitted by: Gad Shaulsky [[log in to unmask]]
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[End dictyNews, volume 42, number 29]
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