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| Status |
Public on Dec 01, 2023 |
| Title |
Transcriptional response to gut pathobiont Streptococcus gallolyticus subsp. gallolyticus infection of huham colon cells |
| Organism |
Homo sapiens |
| Experiment type |
Expression profiling by high throughput sequencing
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| Summary |
Streptococcus gallolyticus sp. gallolyticus (SGG) is a gut pathobiont involved in the development of colorectal cancer (CRC). To decipher SGG contribution in tumor initiation and/or acceleration respectively, a global transcriptome was performed in normal colonic cells (FHC) and in tumoral colonic cells (HT29). To identify SGG-specific alterations, we chose the phylogenetically closest relative, Streptococcus gallolyticus subsp. macedonicus (SGM) as control bacterium. We show that SGM, a bacterium generally considered as safe, did not induce any transcriptional changes on the two human colonic cells FHC and HT29. The transcriptional reprogramming induced by SGG in FHC and HT29 cells was significantly different, however most of the genes up- and down-regulated were associated with cancer disease. Top up-regulated genes related to cancer were: (i) IL-20, CLK1, SORBS2, ERG1, PIM1, SNORD3A for normal FHC cells and (ii) TSLP, BHLHA15, LAMP3, ZNF27B, KRT17, ATF3 for cancerous HT29 cells. SGG induces much stronger transcriptional changes in cancerous than in normal colonic cells (2090 vs genes 128). Gene set enrichment analysis reveals that SGG-induced strong ER- (endoplasmic reticulum) stress and UPR- (unfolded protein response) activation in colonic epithelial cells. Our results suggest that SGG induces a pro-tumoral shift in human colonic cells particularly in transformed cells potentially accelerating tumor development in the colon.
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| Overall design |
FHC cells were seeded into 6-well plates at a density of 2 x 105 cells per well and HT29 cells at 4 x 105 cells per well and incubated for 16-20 hours. Stationary phase bacteria were scratched from fresh THY plates (overnight culture) and washed with sterile phosphate buffered saline, pH 7.4 (PBS) to get an OD600 of 1 (corresponding to 6.5 x 108 CFU). The bacteria were diluted in DMEM (Gibco, Ref. 12320032, low glucose, pyruvate, HEPES) to obtain the desired concentration as follows (i) SGM of 6.5 x 105 CFU/ml and (ii) SGG UCN34 of 6.5 x 104 CFU/ml. Cells are washed once with DMEM and then infected with the medium containing the bacteria by adding 25 ml of bacterial suspension per well in the 6-well plate. For the non-treated (NT) condition, only 2 ml of fresh media was added. For each condition, a complete 6-well plate was used and then pooled together to obtain enough cellular material for further RNA extraction. Trimethoprim (Sigma, Ref. T7883), a bacteriostatic antibiotic was added at 50 μg.ml-1 final concentration after 6 h of co-culture to prevent media acidification due to bacterial growth. The total co-culture incubation time was 24 h. Total RNA was extracted from cell monolayers with the RNeasy Plus Mini kit (Qiagen, USA), according to the manufacturer's instructions. Three independent replicates for each condition were performed and analyzed. RNAs were conserved at -80°C and used in transcriptome Illumina assay and quantitative RT-PCR.
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| Contributor(s) |
Pasquereau-Kotula E, du Merle L, Sismeiro O, Pietrosemoli N, Varet H, Legendre R, Trieu-Cuot P, Dramsi S |
| Citation(s) |
38033043 |
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| Submission date |
May 10, 2023 |
| Last update date |
Dec 02, 2023 |
| Contact name |
Hugo Varet |
| Organization name |
Institut Pasteur
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| Street address |
28 rue du Docteur Roux
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| City |
Paris |
| ZIP/Postal code |
75015 |
| Country |
France |
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| Platforms (1) |
| GPL16791 |
Illumina HiSeq 2500 (Homo sapiens) |
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| Samples (18)
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| Relations |
| BioProject |
PRJNA971227 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSE232211_RAW.tar |
80.7 Mb |
(http)(custom) |
TAR (of TXT) |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
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