Journal: eLife

Article Title: Unique integrated stress response sensors regulate cancer cell susceptibility when Hsp70 activity is compromised

doi: 10.7554/eLife.64977

Figure Lengend Snippet: ( A ) Principal component analysis plot of publicly available RNAseq datasets is presented for tested breast cancer cell lines. MAL3-101-sensitive cells are indicated in aqua green while resistant cell lines are indicated in light red. ( B ) Heatmap representation of significantly differentially expressed proteins between publicly available RPPA datasets for sensitive and resistant cells (MDA MB 231 and MDA MB 453). Wilcoxon signed-rank test between resistant and sensitive cell lines was used to determine proteins that were differentially expressed. p<0.1 was considered significant. ( C ) Principal component analysis plot of RNAseq performed on MAL3-101 treated (Samples A4, A5, A6, B4, B5, B7 ) or DMSO (Samples A1, A2, A7, B1, B2, B3 ) treated sensitive and resistant cells. MDA MB 453 cells (MAL3-101 resistant, sample A) are represented in light red and MDA MB 231 cells (MAL3-101 sensitive, sample B) in aqua green. ( D ) The top 35 differentially expressed genes detected in MAL3-101 treated and untreated cell lines (MDA MB 231 in green and MDA MB 453 in light red) are represented in a heatmap graph. ( E ) Gene Ontology enrichment plot of Biological Processes using significantly differential genes in MDA MB 231 cell lines with and without MAL3-101 treatment. p<0.05 was considered significant.

Article Snippet: To better define the difference in the sensitivities of breast cancer cells to MAL3-101, we analyzed publicly available RNA sequencing (RNAseq) databases from the Broad Institute Cancer Cell Line Encyclopedia (CCLE) ( ) and RNAseq data published in prior work ( ).

Techniques:

Journal: bioRxiv

Article Title: TGFβ signaling in cancer-associated fibroblasts drives a hepatic gp130-dependent pro-metastatic inflammatory program in CMS4 colorectal cancer subtype

doi: 10.1101/2024.06.25.600311

Figure Lengend Snippet: A. Schematic overview of workflow for CAF isolation and TGFβ1 pathway-targeted qPCR array. B. RNA expression of TGFβ1 target genes in two primary CRC CAFs (CAF1 and CAF2) and one LM-CRC CAF. qPCR values have been log transformed. C. Numerical values depicting the seven TGFβ1 target genes that showed consistent upregulation. D. Fold change RNA expression of these seven TGFβ1 target genes (yellow symbols). E. IL6 RNA expression in primary CRC CAFs (CAF4 and CAF5) and the CCD-18Co colon fibroblast line after stimulation with TGFβ1 or medium control. N=3 CCD-18Co, CAF4; N=2 CAF5 independent biological experiments. ** p≤0.01 determined by paired T test. F. Protein expression of IL-6 in tissue lysates of normal colon and primary CRC (left panel, N=50) or normal liver and LM-CRC (right panel, N=50) as determined by ELISA. **** p≤0.0001, * p≤0.05 determined by unpaired T test. G. IL6 RNA expression in unstimulated normal colon fibroblasts (N=8) or primary CRC CAFs (N=10). H. RNA expression of IL6 in the original CMS classified cohort (CMS1: N=457; CMS2: N=1183; CMS3: N=409; CMS4: N=773). **** p≤0.0001 determined by one-way ANOVA with correction for multiple testing (Dunnett’s test).

Article Snippet: IL-6 concentrations in tissue lysates and CAF supernatants were measured using a commercially available kit (Human IL-6 DuoSet ELISA, R&D Systems).

Techniques: Isolation, RNA Expression, Transformation Assay, Control, Expressing, Enzyme-linked Immunosorbent Assay

Journal: bioRxiv

Article Title: TGFβ signaling in cancer-associated fibroblasts drives a hepatic gp130-dependent pro-metastatic inflammatory program in CMS4 colorectal cancer subtype

doi: 10.1101/2024.06.25.600311

Figure Lengend Snippet: A. RNA expression of IL6 in CAFs derived either from the primary tumor (CRC CAF, N=18) or liver metastasis (LM-CRC CAF, N=6). *** p≤0.001 determined by unpaired T test. B. RNA-seq data from GSE46824 dataset showing IL6 RNA expression in an independent cohort of primary CRC CAFs (N=14) and LM-CRC CAFs (N=11). *** p≤0.001 determined by unpaired T test. C. IL-6 protein expression in unstimulated primary CRC CAF (N=8) and LM-CRC CAF (N=6) supernatant determined by ELISA. ** p≤0.01 determined by unpaired T test. D, E. IL-6 protein expression in TGFβ1-stimulated (5 ng/ml) primary CRC CAFs (N=8) ( D ) and LM-CRC CAFs (N=6) ( E ). * p≤0.05 determined by unpaired T test. F. Fold change IL-6 protein expression in supernatant from TGFβ1-stimulated vs. unstimulated primary CRC CAFs (N=8) and LM-CRC CAFs (N=6). ns, p>0.05 determined by unpaired T test. G. Percentage decrease in IL-6 protein expression in supernatant in primary CRC (N=5) and LM-CRC (N=5) CAFs after inhibition with SB431532 (Alk5 inhibitor). ns, p>0.05 determined by unpaired T test. H. RNA-seq data from GSE46824 dataset (2B) showing TGFβ1 RNA expression in an independent cohort of primary CRC CAFs (N=14) and LM-CRC CAFs (N=11). * p≤0.05 determined by unpaired T test. I. TGFβ1 protein expression in unstimulated primary CRC CAF (N=5) and LM-CRC CAF (N=6) supernatant determined by ELISA. * p≤0.05 determined by unpaired T test.

Article Snippet: IL-6 concentrations in tissue lysates and CAF supernatants were measured using a commercially available kit (Human IL-6 DuoSet ELISA, R&D Systems).

Techniques: RNA Expression, Derivative Assay, RNA Sequencing, Expressing, Enzyme-linked Immunosorbent Assay, Inhibition

Journal: bioRxiv

Article Title: TGFβ signaling in cancer-associated fibroblasts drives a hepatic gp130-dependent pro-metastatic inflammatory program in CMS4 colorectal cancer subtype

doi: 10.1101/2024.06.25.600311

Figure Lengend Snippet: A-C. RNA expression of the three TGFβ isoforms in the original CMS cohort (CMS1: N=457; CMS2: N=1183; CMS3: N=409; CMS4: N=773). **** p≤0.0001 determined by one-way ANOVA with correction for multiple testing (Dunnett’s test). D. Protein expression of TGFβ isoforms in the supernatant of primary CRC and LM-CRC CAFs 48 hours after start of serum starvation as determined by ELISA. E, F. Expression of IL-6 in supernatant of primary CRC CAFs (N=7) ( E ) or LM-CRC CAFs (N=6) ( F ) 48 hours after start stimulation with either TGFβ2 or TGFβ3. * p≤0.05 determined by paired T test.

Article Snippet: IL-6 concentrations in tissue lysates and CAF supernatants were measured using a commercially available kit (Human IL-6 DuoSet ELISA, R&D Systems).

Techniques: RNA Expression, Expressing, Enzyme-linked Immunosorbent Assay

Journal: Genome Research

Article Title: Reevaluation of the Toxoplasma gondii and Neospora caninum genomes reveals misassembly, karyotype differences, and chromosomal rearrangements

doi: 10.1101/gr.262832.120

Figure Lengend Snippet: Comparative analysis of genome assemblies of Neospora caninum and Toxoplasma gondii using third-generation sequencing data reveals misassembly and karyotype differences. ( A ) Comparative analysis of the T. gondii type II ( Tg ME49) genome assembly and the N. caninum Liverpool ( Nc Liv) strain genome assembly, obtained based on Sanger technology sequencing data. ( B ) Comparative alignment of the Nc Liv genome assemblies using Sanger and third-generation (long-read) technology. ( C ) Comparative alignment of the T. gondii type II ( Tg ME49) genome assemblies based on Sanger technology sequencing data or third-generation (long-read) technology of T. gondii type I ( Tg RH). ( D ) Comparative alignment of the T. gondii type I ( Tg RH) and the Nc Liv genome assemblies based on third-generation (long-read) sequencing technology. ( E ) Chromosomal layout of N. caninum . Karyotype, chromosome length, telomeres, putative centromeres, and large repeats are shown.

Article Snippet: Nonetheless, examination of available T. gondii PRU RNA sequence data from Oxford Nanopore has revealed numerous reads capable of encoding full-length cytochrome transcripts.

Techniques: Sequencing

Journal: Genome Research

Article Title: Reevaluation of the Toxoplasma gondii and Neospora caninum genomes reveals misassembly, karyotype differences, and chromosomal rearrangements

doi: 10.1101/gr.262832.120

Figure Lengend Snippet: Regions of synteny breaks between N. caninum and T. gondii are populated by three conserved domains. ( A ) Sequence identity of domains identified at regions where chromosomal rearrangements have occurred. ( B ) Graphical representation of Chromosome VIII of Nc Liv. Comparative alignment to the T. gondii chromosomes. Percentages of sequence identity are shown. Regions examined for the presence of motifs are indicated (light green). The position of the putative centromere is indicated in orange. Note that large repetitive regions were not identified in this chromosome. 5′ (light purple) and 3′ (dark purple) telomeres are indicated. The identity and number of domains found per region, in Chromosome VII, are indicated.

Article Snippet: Nonetheless, examination of available T. gondii PRU RNA sequence data from Oxford Nanopore has revealed numerous reads capable of encoding full-length cytochrome transcripts.

Techniques: Sequencing

Journal: Genome Research

Article Title: Reevaluation of the Toxoplasma gondii and Neospora caninum genomes reveals misassembly, karyotype differences, and chromosomal rearrangements

doi: 10.1101/gr.262832.120

Figure Lengend Snippet: Comparative analysis of mitochondrial genome structures and annotations of Neospora and Toxoplasma reveals gene fragmentation and reshuffling between species and strains. ( A ) The repetitive nature of the gene structure in a 32-kb mitochondrial DNA contig of Nc Liv is graphically represented in a YASS plot. ( B ) The repetitive nature of the gene structure in a 16-kb mitochondrial DNA contig of Nc Liv is graphically represented in a YASS plot. ( C ) Comparative alignment between two Nc Liv mitochondrial contigs of 16 and 32 kb, respectively. ( D ) Comparative alignment between a Nc Liv mitochondrial contig of 32 kb and a Nc Uru1 mitochondrial contig of 38 kb. ( E ) Comparative alignment between two Nc Uru1 mitochondrial contigs of 16 and 38 kb, respectively. ( F ) The repetitive nature of the gene structure in a 16-kb mitochondrial DNA contig of Nc Uru1 is graphically represented in a YASS plot. ( G ) Comparative alignment between a Nc Liv mitochondrial contig of 32 kb and a T. gondii mitochondrial contigs of 39 kb.

Article Snippet: Nonetheless, examination of available T. gondii PRU RNA sequence data from Oxford Nanopore has revealed numerous reads capable of encoding full-length cytochrome transcripts.

Techniques:

Journal: bioRxiv

Article Title: Abrogation and Homeostatic Restoration of IgE Responses by a Universal IgE Allergy CTL Vaccine—The Three Signal Self/Non-Self/Self (S/NS/S) Model

doi: 10.1101/2023.10.12.561777

Figure Lengend Snippet: MoDCs were prepared according to the procedures , described in the Suppl. For drug treatment (A-F), MoDCs were incubated with different drugs at the concentrations indicated. The duration of treatment was 4 hours unless otherwise indicated. Whole-cell lysates were prepared after treatment and resolved by SDS‒PAGE followed by immunoblotting using corresponding antibodies (upper panel), and the relative signal was quantified by ImageJ (lower panel). A. AICAR/metformin (Met), CDCA and GSK-J4 downregulate SREBP-1 in immature MoDCs, as shown by immunoblotting (upper panel) and ImageJ (lower panel). B. AICAR/Met, 6ECDCA and VD3 (100 nM calcitriol) downregulate SREBP-1 in mature MoDCs by immunoblotting and ImageJ. C. OEA downregulates SREBP-1 with others as positive control. D . 6ECDCA, GSK-J4 and OEA downregulate FASN in mature MoDCs with 6ECDCA as control. E . AICAR/Met, CDCA, 6ECDCA and OEA upregulate SHP in mature MoDCs. F . AICAR/Met and GSK-J4 upregulate CTLA-4 in mature MoDCs. G . Mature MoDCs were transfected with SREBP- 1c siRNA or control scRNA at a final concentration of 33 nM 24 hours later, and RNA was isolated followed by qRT‒PCR detailed in Supplement. H. Homeostatic lipogenic model for generative DCs, explained in text for three signal model.

Article Snippet: No sequences available siRNA for SREBP-1 knockdown: SREBP-1 siRNA (h) (# sc-36557, Santa Cruz Biotechnology) No sequences available. qRT-PCR primers: SREBP-1 (h) Forward: 5’-ACAGTGACTTCCCTGGCCTAT-3’ Reverse: 5’-GCATGGACGGGTACATCTTCAA-3’ FOXP3 (h) Forward: 5′-CACAACATGCGACCCC CTTTCACC-3′ Reverse: 5′-AGGTTGTGGCGGAT GGCGTTCTTC-3′ GAPDH (h) Forward: 5’-TTG GCT ACA GCA ACA GGGTG-3’ Reverse: 5’-GGG GAG ATT CAG TGT GGT GG-3’

Techniques: Incubation, Western Blot, Positive Control, Control, Transfection, Concentration Assay, Isolation