Coactosin-like protein 1 inhibits neuronal migration during mouse corticogenesis

Coactosin-like protein 1 (Cotl1), a member of the actin-depolymerizing factor (ADF)/cofilin family, was first purified from a soluble fraction of Dictyostelium discoideum cells. Neuronal migration requires cytoskeletal remodeling and actin regulation. Although Cotl1 strongly binds to F-actin, the role of Cotl1 in neuronal migration remains undescribed. In this study, we revealed that Cotl1 overexpression impaired migration of both early- and late-born neurons during mouse corticogenesis. Moreover, Cotl1 overexpression delayed, rather than blocked, neuronal migration in late-born neurons. Cotl1 expression disturbed the morphology of migrating neurons, lengthening the leading processes. This study is the first to investigate the function of Cotl1, and the results indicate that Cotl1 is involved in the regulation of neuronal migration and morphogenesis.

Analysis of static and dynamic balance in healthy elderly practitioners of Tai Chi Chuan versus ballroom dancing

OBJECTIVE: To determine whether Tai Chi Chuan or ballroom dancing promotes better performance with respect to postural balance, gait, and postural transfer among elderly people. METHODS: We evaluated 76 elderly individuals who were divided into two groups: the Tai Chi Chuan Group and the Dance Group. The subjects were tested using the NeuroCom Balance Master¯ force platform system with the following protocols: static balance tests (the Modified Clinical Tests of Sensory Interaction on Balance and Unilateral Stance) and dynamic balance tests (the Walk Across Test and Sit-to-stand Transfer Test). RESULTS: In the Modified Clinical Test of Sensory Interaction on Balance, the Tai Chi Chuan Group presented a lower sway velocity on a firm surface with open and closed eyes, as well as on a foam surface with closed eyes. In the Modified Clinical Test of Sensory Interaction on Unilateral Stance, the Tai Chi Chuan Group presented a lower sway velocity with open eyes, whereas the Dance Group presented a lower sway velocity with closed eyes. In the Walk Across Test, the Tai Chi Chuan Group presented faster walking speeds than those of the Dance Group. In the Sit-to-stand Transfer Test, the Tai Chi Chuan Group presented shorter transfer times from the sitting to the standing position, with less sway in the final standing position. CONCLUSION: The elderly individuals who practiced Tai Chi Chuan had better bilateral balance with eyes open on both types of surfaces compared with the Dance Group. The Dance Group had better unilateral postural balance with eyes closed. The Tai Chi Chuan Group had faster walking speeds, shorter transfer times, and better postural balance in the final standing position during the Sit-to-stand Test. .

Analysis of rapamycin induced autophagy in Dictyostelium discoideum.

Natural autophagy and autophagic cell death is being studied in the model system, D. discoideum, which has well known genetic and experimental advantages over the other known systems. There is no apoptotic machinery present in this organism which could interfere with the non-apoptotic cell death. The target of rapamycin (TOR) pathway is a major nutrient-sensing pathway which when inhibited by the drug rapamycin induces autophagy. Rapamycin was originally discovered as an anti-fungal agent but its use was abandoned when it was discovered to have potent immunosuppressive and anti-proliferative properties. It is a known drug used today for various cancer treatments and also for increasing longevity in many model organisms. It has a wide usage but its effects on other pathways or molecules are not known. This model system was used to study the action of rapamycin on autophagy induction. Using the GFP-Atg8, an autophagosome marker, it was shown that rapamycin treatment can induce autophagy by an accumulation of reactive oxygen species and intracellular free calcium. Rapamycin suppresses proliferation by induction of cell cycle arrest in the G1 phase. Taken together, the results suggest that the core machinery for autophagy is conserved in D. discoideum and it can serve as a good model system to delineate the action of rapamycin induced autophagy.

Peptide: N- glycanase is expressed in prestalk cells and plays a role in the differentiation of prespore cells during development of Dictyostelium discoideum.

Peptide: N- glycanase (PNGase) enzyme is found throughout eukaryotes and plays an important role in the misfolded glycoprotein degradation pathway. This communication reports the expression patterns of the pngase transcript (as studied by the analysis of β- galactosidase reporter driven by the putative pngase promoter) and protein (as studied by the analysis of β- galactosidase reporter expressed under the putative pngase promoter as a fusion with the pngase ORF) during development and further elucidated the developmental defects of the cells lacking PNGase (png-). The results show that the DdPNGase is an essential protein expressed throughout development and β- galactosidase activity was present in the anterior part of the slug. In structures derived from a null mutant for pngase, the prestalk A and AO patterning was expanded and covered a large section of the prespore region of the slugs. When developed as chimeras with wild type, the png- cells preferentially populate the prestalk/stalk region. When the mutants were mixed in higher ratios, they also tend to form the prespore/spore cells. The results emphasize that the DdPNGase has an essential role during development and the mutants have defects in a system that changes the physiological dynamics in the prespore cells. DdPNGase play a role in development both during aggregation and in the differentiation of prespore cells.

Spliceosomal genes in the D. discoideum genome: a comparison with those in H. sapiens, D. melanogaster, A. thaliana and S. cerevisiae

Little is known about pre-mRNA splicing in Dictyostelium discoideum although its genome has been completely sequenced. Our analysis suggests that pre-mRNA splicing plays an important role in D. discoideum gene expression as two thirds of its genes contain at least one intron. Ongoing curation of the genome to date has revealed 40 genes in D. discoideum with clear evidence of alternative splicing, supporting the existence of alternative splicing in this unicellular organism. We identified 160 candidate U2-type spliceosomal proteins and related factors in D. discoideum based on 264 known human genes involved in splicing. Spliceosomal small ribonucleoproteins (snRNPs), PRP19 complex proteins and late-acting proteins are highly conserved in D. discoideum and throughout the metazoa. In non-snRNP and hnRNP families, D. discoideum orthologs are closer to those in A. thaliana, D. melanogaster and H. sapiens than to their counterparts in S. cerevisiae. Several splicing regulators, including SR proteins and CUG-binding proteins, were found in D. discoideum, but not in yeast. Our comprehensive catalog of spliceosomal proteins provides useful information for future studies of splicing in D. discoideum where the efficient genetic and biochemical manipulation will also further our general understanding of pre-mRNA splicing.

Arachidonic acid is a chemoattractant for Dictyostelium discoideum cells.

Cyclic AMP (cAMP)is a natural chemoattractant of the social amoeba Dictyostelium discoideum. It is detected by cell surface cAMP receptors. Besides a signalling cascade involving phosphatidylinositol 3,4,5-trisphosphate (PIP3), Ca2+ signalling has been shown to have a major role in chemotaxis. Previously, we have shown that arachidonic acid (AA) induces an increase in the cytosolic Ca2+ concentration by causing the release of Ca2+ from intracellular stores and activating influx of extracellular Ca2+. Here we report that AA is a chemoattractant for D. discoideum cells differentiated for 8-9 h. Motility towards a glass capillary filled with an AA solution was dose-dependent and qualitatively comparable to cAMP-induced chemotaxis. Ca2+ played an important role in AA chemotaxis of wild-type Ax2 as ethyleneglycol-bis(b-aminoethyl)-N,N,N',N'-tetraacetic acid (EGTA) added to the extracellular buffer strongly inhibited motility. In the HM1049 mutant whose iplA gene encoding a putative Ins(1,4,5)P3 -receptor had been knocked out, chemotaxis was only slightly affected by EGTA. Chemotaxis in the presence of extracellular Ca2+ was similar in both strains. Unlike cAMP, addition of AA to a cell suspension did not change cAMP or cGMP levels. A model for AA chemotaxis based on the findings in this and previous work is presented.

A model for cell type localization in the migrating slug of Dictyostelium discoideum based on differential chemotactic sensitivity to cAMP and differential sensitivity to suppression of chemotaxis by ammonia.

The three basic cell types in the migrating slug of Dictyostelium discoideum show differential chemotactic response to cyclic AMP (cAMP) and differential sensitivity to suppression of the chemotaxis by ammonia.The values of these parameters indicate a progressive maturation of chemotactic properties during the transdifferentiation of slug cell types.We present a model that explains the localization of the three cell types within the slug based on these chemotactic differences and on the maturation of their chemotactic properties.

Signaling molecules involved in the transition of growth to development of Dictyostelium discoideum.

The social amoeba Dictyostelium discoideum, a powerful paradigm provides clear insights into the regulation of growth and development. In addition to possessing complex individual cellular functions like a unicellular eukaryote, D. discoideum cells face the challenge of multicellular development. D. discoideum undergoes a relatively simple differentiation process mainly by cAMP mediated pathway. Despite this relative simplicity, the regulatory signaling pathways are as complex as those seen in metazoan development. However, the introduction of restriction-enzyme-mediated integration (REMI) technique to produce developmental gene knockouts has provided novel insights into the discovery of signaling molecules and their role in D. discoideum development. Cell cycle phase is an important aspect for differentiation of D. discoideum, as cells must reach a specific stage to enter into developmental phase and specific cell cycle regulators are involved in arresting growth phase genes and inducing the developmental genes. In this review, we present an overview of the signaling molecules involved in the regulation of growth to differentiation transition (GDT), molecular mechanism of early developmental events leading to generation of cAMP signal and components of cAMP relay system that operate in this paradigm.

Cellular automata approach to biological pattern formation. (II): The growth pattern of bacterial colonies / 生物医学工程学杂志

The investigation of the mechanism of biological pattern has been an important topic of life sciences, especially of developmental biology, for a long time. We have established a cellular automata model of biological pattern formation, which defines how to form a biological pattern from individual cells and their behaviors, cell-cell interactions, and cell-environment interactions. In this paper, we use that model to simulate the growth pattern of the bacterial colonies in consideration of the effects of both nutrient and metabolite on the cell's reproduction.

Cellular automata approach to biological pattern formation (I): the aggregation pattern in dictyostelium discoideum / 生物医学工程学杂志

The investigation of the mechanism of biological pattern has been an important topic of life sciences, especially, of developmental biology, for a long time. It is an interdisciplinary problem and many researching data have been obtained and some theories have been structured from many points of view in science. However, up to now, the actual mechanism is still a fascinating puzzle and needs more studies. In this paper, we try to construct a cellular automata model of biological pattern. This model defines the individual model cells and their behaviors, cell-cell interactions, and cell-environment interactions. As an application, we present a new discrete model to simulate the aggregation phase of the development of Dictyostelium discoideum with the concept of "inducing switch".

Expression of the soluble human Fas ligand in Dictyostelium discoideum / 生物工程学报

An expression system is described for high-yield production of recombinant soluble human FasL (shFasL) in Dictyostelium discoideum cells. DNA encoding amino acids 141 - 281 of hFasL was PCR amplified from cDNA derived from activated human neutrophils. The resulting product was fused with a DNA fragment encoding hCG-beta signal peptide and cloned in the expression vector pMB12neo. Dictyostelium strain AX3 was transfected with this plasmid, yielding a recombinant strain called AX3-pCESFL95-H3. In order to improve the shFasL expression level, pMB12neo was optimized by replacing its transcriptional terminator/ polyadenylation segment of the 2H3 gene with an actin8 terminator/polyadenylation segment, yielding derived expression vector pMB74. The recombinant Dictyostelium strain called AX3-pLu8 was generated with this new plasmid. When the recombinant cells were cultivated in a complex HL-5C medium, a cell density of (1.5 - 2) x 10(7)/mL was reached, and the shFasL level expressed by strains AX3-pCESFL95-H3 and AX3-pLu8 was 23.5 microg/L and 206 microg/L, respectively. By using a newly developed synthetic medium called SIH as culture medium, higher cell density of (4 - 5) x 10(7)/mL was achieved. Correspondently, 111 microg/L and 420 microg/L shFasL were secreted by recombinant strains AX3-pCESFL95-H3 and AX3-pLu8, respectively.

Calcium regulates the expression of a Dictyostelium discoideum asparaginyl tRNA synthetase gene.

In a screen for calcium-regulated gene expression during growth and development of Dictyostelium discoideum we have identified an asparaginyl tRNA synthetase (ddAsnRS) gene, the second tRNA synthetase gene identified in this organism. The ddAsnRS gene shows many unique features. One, it is repressed by lowering cellular calcium, making it the first known calcium-regulated tRNA synthetase. Two, despite the calcium-dependence, its expression is unaltered during the cell cycle, making this the first D. discoideum gene to show a calcium-dependent but cell cycle phase-independent expression. Finally, the N-terminal domain of the predicted ddAsnRS protein shows higher sequence similarity to Glutaminyl tRNA synthetases than to other Asn tRNA synthetases. These unique features of the AsnRS from this primitive eukaryote not only point to a novel mechanism regulating the components of translation machinery and gene expression by calcium, but also hint at a link between the evolution of GlnRS and AsnRS in eukaryotes.

Effect of arsenic on cell growth of the cellular slime mould, Dictyostelium discoideum.

The growing D. discoideum cells were killed in a dose-dependent manner when exposed to 100 and 140 ppm of arsenic (As2O3) at mid-log phase for 20 min. Reduced plaque sizes and changed cell and colony morphologies were observed in the treated cells. Endocytotic functions (both phagocytosis and pinocytosis) were also inhibited in the treated cells. Arsenic treated cell showed a lower DNA and protein synthetic activities. These findings are discussed in relation to known mechanism of action of the heavy metal on growth-related cellular functions.

Biochemical basis of the high resistance to oxidative stress in Dictyostelium discoideum.

Aerobic organisms experience oxidative stress due to generation of reactive oxygen species during normal aerobic metabolism. In addition, several chemicals also generate reactive oxygen species which induce oxidative stress. Thus oxidative stress constitutes a major threat to organisms living in aerobic environments. Programmed cell death or apoptosis is a physiological mechanism of cell death, that probably evolved with multicellularity, and is indispensable for normal growth and development. Dictyostelium discoideum, an eukaryotic developmental model, shows both unicellular and multicellular forms in its life cycle and exhibits apparent caspase-independent programmed cell death, and also shows high resistance to oxidative stress. An attempt has been made to investigate the biochemical basis for high resistance of D. discoideum cell death induced by different oxidants. Dose-dependent induction of cell death by exogenous addition of hydrogen peroxide (H2O2), in situ generation of H2O2 by hydroxylamine, and nitric oxide (NO) generation by sodium nitroprusside treatment in D. discoideum were studied. The AD50 doses (concentration of the oxidants causing 50% of the cells to die) after 24 h of treatment were found to be 0.45 mM, 4 mM and 1 mM, respectively. Studies on enzymatic antioxidant status of D. discoideum when subjected to oxidative stress, NO and nutrient stress reveal that superoxide dismutase and catalase were unchanged; a significant induction of glutathione peroxidase was observed. Interestingly, oxidative stress-induced lipid membrane peroxidative damage could not be detected. The results shed light on the biochemical basis for the observed high resistance to oxidative stress in D. discoideum.

On the origin of differentiation.

Following the origin of multicellularity in many groups of primitive organisms there evolved more than one cell type. It has been assumed that this early differentiation is related to size the larger the organism the more cell types. Here two very different kinds of organisms are considered: the volvocine algae that become multicellular by growth, and the cellular slime moulds that become multicellular by aggregation. In both cases there are species that have only one cell type and others that have two. It has been possible to show that there is a perfect correlation with size: the forms with two cell types are significantly larger than those with one. Also in both groups there are forms of intermediate size that will vary from one to two cell types depending on the size of the individuals, suggesting a form of quorum sensing. These observations reinforce the view that size plays a critical role in influencing the degree of differentiation.

Mining microorganism EST databases in the quest for new proteins

Microorganisms with large genomes are commonly the subjects of single-round partial sequencing of cDNA, generating expressed sequence tags (ESTs). Usually there is a great distance between gene discovery by EST projects and submission of amino acid sequences to public databases. We analyzed the relationship between available ESTs and protein sequences and used the sequences available in the secondary database, clusters of orthologous groups (COG), to investigate ESTs from eight microorganisms of medical and/or economic relevance, selecting for candidate ESTs that may be further pursued for protein characterization. The organisms chosen were Paracoccidioides brasiliensis, Dictyostelium discoideum, Fusarium graminearum, Plasmodium yoelii, Magnaporthe grisea, Emericella nidulans, Chlamydomonas reinhardtii and Eimeria tenella, which have more than 10,000 ESTs available in dbEST. A total of 77,114 protein sequences from COG were used, corresponding to 3,201 distinct genes. At least 212 of these were capable of identifying candidate ESTs for further studies (E. tenella). This number was extended to over 700 candidate ESTs (C. reinhardtii, F. graminearum). Remarkably, even the organism that presents the highest number of ESTs corresponding to known proteins, P. yoelii, showed a considerable number of candidate ESTs for protein characterization (477). For some organisms, such as P. brasiliensis, M. grisea and F. graminearum, bioinformatics has allowed for automatic annotation of up to about 20 of the ESTs that did not correspond to proteins already characterized in the organism. In conclusion, 4093 ESTs from these eight organisms that are homologous to COG genes were selected as candidates for protein characterization

Identificação de genes codificadores de serina/treonina fosfatases em Dictyostelium discoideum e caracterização funcional da proteína fosfatase do tipo 4 (PP4) / Identification of phosphatases serine/treonine genes coders in Dictyostelium discoideum and functional characterization of phosphatases protein type 4 (PP4)

As serina/treonina fosfatases (PPs) são enzimas responsáveis pela desfosforilação de resíduos de fosfoserina e/ou fosfotreonina e estão subdivididas em duas famílias gênicas designadas PPP e PPM. A família PPP está dividida em cinco subfamílias, que compreendem as PPs do tipo 1 (PP1), 2A (PP2A), 2B (PP2B), 5 (PP5) e 7 (PP7), de acordo com a similaridade entre as sequências de aminoácidos das subunidades ou domínios destas enzimas. Novas PPs estão sendo descobertas em diferentes organismos e classificadas nestas famílias ou subfamílias com base na análise comparada de suas sequências. Uma delas é a proteína fosfatase do tipo 4 (PP4), descoberta originalmente em coelhos e cujas funções biológicas vêm sendo progressivamente elucidadas...