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| Status |
Public on Nov 27, 2013 |
| Title |
Reversible mRNA and miRNA expression patterns in the transcriptome of Rasless fibroblasts |
| Platform organisms |
Mus musculus; synthetic construct |
| Sample organism |
Mus musculus |
| Experiment type |
Expression profiling by array
Non-coding RNA profiling by array
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| Summary |
Analysis of the transcriptional profiles of mRNA and microRNA in Rasless fibroblasts. 4-Hydroxy-tamoxifen (4-OHT) treatment triggers removal of K-Ras expression in [H-Ras-/-;N-Ras-/-;K-Raslox/lox;RERTert/ert ] mouse fibroblasts (named K-Raslox) generating “Rasless” MEFs which are unable to proliferate, but recover proliferative ability after ectopic expression of constitutively active downstream kinases such as BRAF and MEK1.
Using Affymetrix microarrays, we compared the transcriptional profiles of Rasless fibroblasts to those of K-Raslox (MEFs lacking only H-Ras and N-Ras). We also compared the transcriptional profile of Rasless cells with those of BRAF and MEK1-transfected Rasless cells which recovered proliferative ability. Re-entry of the Rasless cells into cell cycle progression through ectopic expression of activated BRAF or MEK1 resulted in full reversion of most transcriptional alterations identified in cells lacking the three canonical Ras proteins.
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| Overall design |
Pre-confluent cell cultures corresponding to the different Ras genotypes were harvested for extraction of total RNA and miRNA samples, and subsequent hybridization on Affymetrix microarrays. The different sets of experimental mRNA samples analyzed here included (i) 8 samples K-Raslox (proliferating cells expressing only K-Ras, considered as Control): 4 biological replicates, technical duplicates each; (ii) 7 samples Rasless: the same cells after treatment with 4-OHT for 12 days (to become non-proliferating fibroblasts), including 4 biological replicates and technical duplicates from 3 of them; (iii) 3 samples BRAF-rescued MEFs (Rasless cells harboring activated BRAF constructs, with regained proliferative ability): 3 biological replicates; (iv) 3 samples MEK1-rescued MEFs (Rasless cells harboring activated MEK1 constructs, with regained proliferative ability): 3 biological replicates; and their respective controls harbouring the vempty vectors (v) 2 samples Hygromycin-resistant (control for BRAF-transfected Rasless cells) including 2 biological replicates, and (vi) 4 samples Puromycin-resistant (control for MEK1-transfected Rasless cells) including 2 biological replicates, technical duplicates each. The gene expression study contains a total of 27 samples.
Regarding the miRNA expression study, the experimental samples analyzed included (i) 4 samples samples K-Raslox (used as Control): 2 biological replicates, technical duplicates each; (ii) 8 samples Rasless: the same cells after treatment with 4-OHT for 6 days (2 biological replicates, technical duplicates each) and 12 days (2 biological replicates, technical duplicates each); (iii) 4 samples BRAF-rescued, including 2 biological replicates, technical duplicates each; (iv) 4 samples MEK1-rescued,including 2 biological replicates, technical duplicates each, and (v) 4 samples Puromycin-resistant empty vector, containing 2 biological replicates, technical duplicates each. The miRNA expression study contains a total of 24 samples.
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| Contributor(s) |
Azrak S, Ginel-Picardo A, Santos E |
| Citation(s) |
24156637 |
| Submission date |
Mar 15, 2013 |
| Last update date |
Feb 11, 2019 |
| Contact name |
Alicia Ginel Picardo |
| E-mail(s) |
algi@usal.es
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| Phone |
923294801
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| Fax |
923294743
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| Organization name |
Cic, Salamanca
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| Lab |
Laboratory 1
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| Street address |
Campus Miguel de Unamuno
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| City |
Salamanca |
| State/province |
Select a State or Province |
| ZIP/Postal code |
37007 |
| Country |
Spain |
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| Platforms (2) |
| GPL1261 |
[Mouse430_2] Affymetrix Mouse Genome 430 2.0 Array |
| GPL8786 |
[miRNA-1] Affymetrix Multispecies miRNA-1 Array |
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| Samples (51)
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| GSM1099338 |
DU315-6-S.r. (K-Raslox), technical rep2 |
| GSM1099339 |
DU315-6-N (K-Raslox), biological rep3 |
| GSM1099340 |
DU315-6-N2 (K-Raslox), technical rep3 |
| GSM1099341 |
DU244-1-N (K-Raslox). biological rep4 |
| GSM1099342 |
DU244-1-N2 (K-Raslox). technical rep4 |
| GSM1099343 |
DU315-6-12d-4OHT-M (Rasless), biological rep1 |
| GSM1099344 |
DU315-6-12d-4OHT-M.r. (Rasless), technical rep1 |
| GSM1099345 |
DU315-6-12d-4OHT-S (Rasless), biological rep2 |
| GSM1099346 |
DU315-6-12d-4OHT-S.r. (Rasless), technical rep2 |
| GSM1099347 |
DU315-6-12d-4OHT-N (Rasless), biological rep3 |
| GSM1099348 |
DU244-1-12d-4OHT-N (Rasless), biological rep4 |
| GSM1099349 |
DU244-1-12d-4OHT-N.r. (Rasless), technical rep4 |
| GSM1099350 |
LG7-6-5 (BRAF-transfected), biological rep1 |
| GSM1099351 |
LG7-6-9 (BRAF-transfected), biological rep2 |
| GSM1099352 |
LG7-6-13 (BRAF-transfected), biological rep3 |
| GSM1099353 |
MCL1-6-14 (MEK1-transfected), biological rep1 |
| GSM1099354 |
MCL1-6-18 (MEK1-transfected), biological rep2 |
| GSM1099355 |
MCL1-6-25 (MEK1-transfected), biological rep3 |
| GSM1099356 |
MCL23-1-5 (Puromycin-resistant empty vector), biological rep1 |
| GSM1099357 |
MCL23-1-5.r. (Puromycin-resistant empty vector), technical rep1 |
| GSM1099358 |
MCL23-1-2 (Puromycin-resistant empty vector), biological rep2 |
| GSM1099359 |
MCL23-1-2.r. (Puromycin-resistant empty vector), technical rep2 |
| GSM1099360 |
JU10-2-2 (Hygromycin-resistant empty vector), biological rep1 |
| GSM1099361 |
JU10-2-3 (Hygromycin-resistant empty vector), biological rep2 |
| GSM1099362 |
DU315-6 (K-Raslox), biological rep1 |
| GSM1099363 |
DU315-6 (K-Raslox), technical rep1 |
| GSM1099364 |
DU244-1 (K-Raslox). biological rep2 |
| GSM1099365 |
DU244-1 (K-Raslox). technical rep2 |
| GSM1099366 |
DU315-6-6d-4OHT (Rasless), biological rep1 |
| GSM1099367 |
DU315-6-6d-4OHT (Rasless), technical rep1 |
| GSM1099368 |
DU244-1-6d-4OHT (Rasless), biological rep2 |
| GSM1099369 |
DU244-1-6d-4OHT (Rasless), technical rep2 |
| GSM1099370 |
DU315-6-12d-4OHT (Rasless), biological rep1 |
| GSM1099371 |
DU315-6-12d-4OHT (Rasless), technical rep1 |
| GSM1099372 |
DU244-1-12d-4OHT (Rasless), biological rep2 |
| GSM1099373 |
DU244-1-12d-4OHT (Rasless), technical rep2 |
| GSM1099374 |
LG7-6-5 (BRAF-transfected), biological rep1 |
| GSM1099375 |
LG7-6-5 (BRAF-transfected), technical rep1 |
| GSM1099376 |
LG7-6-9 (BRAF-transfected), biological rep2 |
| GSM1099377 |
LG7-6-9 (BRAF-transfected), technical rep2 |
| GSM1099378 |
MCL1-6-18 (MEK1-transfected), biological rep1 |
| GSM1099379 |
MCL1-6-18 (MEK1-transfected), technical rep1 |
| GSM1099380 |
MCL1-6-25 (MEK1-transfected), biological rep2 |
| GSM1099381 |
MCL1-6-25 (MEK1-transfected), technical rep2 |
| GSM1099382 |
MCL23-1-5 (Puromycin-resistant empty vector), biological rep1 |
| GSM1099383 |
MCL23-1-5 (Puromycin-resistant empty vector), technical rep1 |
| GSM1099384 |
MCL23-1-2 (Puromycin-resistant empty vector), biological rep2 |
| GSM1099385 |
MCL23-1-2 (Puromycin-resistant empty vector), technical rep2 |
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| Relations |
| BioProject |
PRJNA193231 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSE45222_RAW.tar |
102.4 Mb |
(http)(custom) |
TAR (of CEL) |
| Processed data included within Sample table |
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