STATc is a key regulator of the transcriptional response to hyperosmotic shock-5

<p><b>Copyright information:</b></p><p>Taken from "STATc is a key regulator of the transcriptional response to hyperosmotic shock"</p><p>http://www.biomedcentral.com/1471-2164/8/123</p><p>BMC Genomics 2007;8():123-123.</p><p>Published online 21 May 2007</p><p>PMCID:PMC1888708.</p><p></p>t activated in cells in response to hypertonicity. In contrast to yeast no osmosensors are so far known in . The OP1 (smostress-dependent athway ) pathway is under control of the hybrid histidine kinase DokA and leads to elevated cAMP levels thereby activating protein kinase A (PKA). The cGMP pathway, which we named OSP (smostress-dependent TATc athway), leads to the activation and nuclear translocation of STATc. Other components of this pathway are probably Rap1, a guanylate cyclase (GC), GbpC, PkyA and PTP3. Based on differential regulation of putative MAPK components we propose a third signalling branch, which is under the control of a MAPK cascade, similar to yeast and mammals. Genes that were up-regulated in our experiments are in red. PM: plasma membrane; NM: nuclear membrane

STATc is a key regulator of the transcriptional response to hyperosmotic shock-3

<p><b>Copyright information:</b></p><p>Taken from "STATc is a key regulator of the transcriptional response to hyperosmotic shock"</p><p>http://www.biomedcentral.com/1471-2164/8/123</p><p>BMC Genomics 2007;8():123-123.</p><p>Published online 21 May 2007</p><p>PMCID:PMC1888708.</p><p></p> if STATc is not involved in the transcriptional regulation. (B) Expected overlap in experiments I and II if STATc is the only transcriptional regulator in response to hypertonicity. (C) Venn diagram of the observed differentially regulated genes from the three comparisons: wt cells treated versus untreated (I), RIC cells treated versus STATctreated (II) and STATctreated versus untreated (III). The numbers of up- and down-regulated genes of the single experiments are printed in red and green, respectively. Genes shared between 2 or 3 comparisons (shaded region) were applied to further analysis

STATc is a key regulator of the transcriptional response to hyperosmotic shock-1

<p><b>Copyright information:</b></p><p>Taken from "STATc is a key regulator of the transcriptional response to hyperosmotic shock"</p><p>http://www.biomedcentral.com/1471-2164/8/123</p><p>BMC Genomics 2007;8():123-123.</p><p>Published online 21 May 2007</p><p>PMCID:PMC1888708.</p><p></p>ts. The data are expressed as means of fold change ± SD of three independent experiments. The corresponding DDB IDs from left to right as follows: DDB0188166, DDB0235172, DDB0188166, DDB0190245, DDB0185120

STATc is a key regulator of the transcriptional response to hyperosmotic shock-4

<p><b>Copyright information:</b></p><p>Taken from "STATc is a key regulator of the transcriptional response to hyperosmotic shock"</p><p>http://www.biomedcentral.com/1471-2164/8/123</p><p>BMC Genomics 2007;8():123-123.</p><p>Published online 21 May 2007</p><p>PMCID:PMC1888708.</p><p></p>ters (1–8) can be distinguished of which clusters 4 and 7 contain those genes that are solely regulated by STATc. The dendrogram is displayed on the left. The differentially regulated genes are depicted as coloured lines. The colour represents the fold of induction (red) or repression (blue) (colour scale see Fig. 3). Non-regulated genes are displayed in yellow. OP1: Osmostress induced pathway 1; OSP: Osmostress induced STATc pathway; SP: STATc pathway irrespective of osmostress. (B) GO biological process terms enriched in cluster 4 and 7. GO tree levels are shown on the left. Bar lengths represent the fold enrichment (scale x-axis). The table indicates the number of genes with a particular annotation in the cluster (List), on the entire array (Total), the significance for enrichment (P-value) and the annotation

STATc is a key regulator of the transcriptional response to hyperosmotic shock-0

<p><b>Copyright information:</b></p><p>Taken from "STATc is a key regulator of the transcriptional response to hyperosmotic shock"</p><p>http://www.biomedcentral.com/1471-2164/8/123</p><p>BMC Genomics 2007;8():123-123.</p><p>Published online 21 May 2007</p><p>PMCID:PMC1888708.</p><p></p>k. After starvation for 4 hours, untreated cells and cells treated for 5 min with 50, 100, 200 and 400 mM sorbitol were fixed with ice cold methanol, and then stained with a monoclonal antibody specific for actin, followed by the incubation with anti-mouse IgG antibody conjugated with Cy5. Size bar is 10 μm. (B) Decrease of cell volume in response to hyperosmotic condition. cells were treated with increasing sorbitol concentrations for 5 minutes. Cell volume was measured with a microcapillary. Values represent the mean of three independent experiments ± standard deviation (SD). (C) Cell survival in response to hyperosmotic shock. Cell survival was measured by plating out treated or untreated cells on lawns and counting the plaques after 2 days of incubation at 21°C. Values represent the mean of three independent experiments ± SD

STATc is a key regulator of the transcriptional response to hyperosmotic shock-2

<p><b>Copyright information:</b></p><p>Taken from "STATc is a key regulator of the transcriptional response to hyperosmotic shock"</p><p>http://www.biomedcentral.com/1471-2164/8/123</p><p>BMC Genomics 2007;8():123-123.</p><p>Published online 21 May 2007</p><p>PMCID:PMC1888708.</p><p></p>1.5 fold in the time course of sorbitol treatment was clustered with GeneSpring 7.2. Four major clusters (1–4) can be distinguished. The dendrogram is displayed on the left. The differentially regulated genes are depicted as coloured lines and the time of treatment in minutes is shown at the bottom. The colour represents the fold induction (red) or repression (blue) as shown in the colour scale below the figure. Non-regulated genes are displayed in yellow. (B) A selection of the GO biological process terms that were enriched in each of the clusters is presented. GO tree levels are shown on the left. Bar lengths represent the fold enrichment (scale x-axis). The table indicates the number of genes with a particular annotation in the cluster (List), on the entire array (Total), the significance for enrichment (P-value) and the annotation. (C) Expression profiles of selected genes from each cluster. The following abbreviations for differentially regulated genes are used. Cluster 1: A, A; B6, 20S proteasome subunit beta-6; C4, 26S proteasome subunit ATPase 4; 11, 26S proteasome non-ATPase regulatory subunit 11; 2, 26S proteasome regulatory subunit 2; C, Cysteine Protease Inhibitor; A, cysteine proteinase 1. Cluster 2: A, coronin; B, actin related protein 2; B, profilin II; A, NCK-Associated Protein; C, actin binding protein; act8, actin8. Cluster 3: C, STATc; A, RasGTPase-activating protein; A, Severin kinase; G21, ABC transporter G family protein. Cluster 4: , vacuolar H+-ATPase subunit

Additional file 1: Table S1. of A set of genes conserved in sequence and expression traces back the establishment of multicellularity in social amoebae

Gene set: The genes identified and additional information on orthology, detection method, and expression peak. (XLSX 97 kb

Image_1_Domain Organization of the UBX Domain Containing Protein 9 and Analysis of Its Interactions With the Homohexameric AAA + ATPase p97 (Valosin-Containing Protein).jpg

The abundant homohexameric AAA + ATPase p97 (also known as valosin-containing protein, VCP) is highly conserved from Dictyostelium discoideum to human and a pivotal factor of cellular protein homeostasis as it catalyzes the unfolding of proteins. Owing to its fundamental function in protein quality control pathways, it is regulated by more than 30 cofactors, including the UBXD protein family, whose members all carry an Ubiquitin Regulatory X (UBX) domain that enables binding to p97. One member of this latter protein family is the largely uncharacterized UBX domain containing protein 9 (UBXD9). Here, we analyzed protein-protein interactions of D. discoideum UBXD9 with p97 using a series of N- and C-terminal truncation constructs and probed the UBXD9 interactome in D. discoideum. Pull-down assays revealed that the UBX domain (amino acids 384–466) is necessary and sufficient for p97 interactions and that the N-terminal extension of the UBX domain, which folds into a β0-α–1-α0 lariat structure, is required for the dissociation of p97 hexamers. Functionally, this finding is reflected by strongly reduced ATPase activity of p97 upon addition of full length UBXD9 or UBXD9261–573. Results from Blue Native PAGE as well as structural model prediction suggest that hexamers of UBXD9 or UBXD9261–573 interact with p97 hexamers and disrupt the p97 subunit interactions via insertion of a helical lariat structure, presumably by destabilizing the p97 D1:D1’ intermolecular interface. We thus propose that UBXD9 regulates p97 activity in vivo by shifting the quaternary structure equilibrium from hexamers to monomers. Using three independent approaches, we further identified novel interaction partners of UBXD9, including glutamine synthetase type III as well as several actin-binding proteins. These findings suggest a role of UBXD9 in the organization of the actin cytoskeleton, and are in line with the hypothesized oligomerization-dependent mechanism of p97 regulation.</p

Additional file 3: of A set of genes conserved in sequence and expression traces back the establishment of multicellularity in social amoebae

Supplementary Figures 1-6. (DOCX 3657 kb

Additional file 2: Table S3. of A set of genes conserved in sequence and expression traces back the establishment of multicellularity in social amoebae

Oligonucleotides primer sequences for knock out studies. (DOC 56 kb