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| Status |
Public on Dec 09, 2008 |
| Title |
Gene expression profiling induced by overexpression and silencing of Ankrd2 |
| Organism |
Mus musculus |
| Experiment type |
Expression profiling by array
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| Summary |
We have used a genomic approach based on microarray technology in order to identify the gene networks that are disturbed by Ankrd2 overexpression and silencing. We performed the experiments at the myoblast stage, because there Ankrd2 localizes in the nucleus where it may act as myogenic transcriptional regulator. The stable alteration of Ankrd2 expression level resulted in the modulation of different muscle differentiation-related genes belonging to functional classes having key roles in myogenesis (cell cycle regulation and apoptosis; skeletal muscle development and contraction; cell adhesion and cytoskeletal organization). The transcriptional profile of Ankrd2-overexpressing clones highlights a remarkable impairment of the myogenic differentiation program. In the same clones the alteration of genes regulating cell cycle and apoptosis shows that the Ankrd2 overexpression leads to raised C2C12 proliferation compromising the response to serum deprivation and then myotube formation. On the other side, in Ankrd2-silenced clones numerous genes promoting cell cycle arrest are found upregulated.
Keywords: gene-specific overexpression and silencing
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| Overall design |
Gene expression profiling experiments have been performed using a mouse microarray platform containing a collection of 13,443 70mer oligonucleotide probes (Qiagen-Operon, version 1.1). Each oligonucleotide was spotted in two replicates on MICROMAX SuperChip I glass slides (Perkin-Elmer) using Biorobotics Microgrid II (Apogent Discoveries), supplied with 48 stealth micro pins.Total RNA was purified from two C2C12 Ankrd2-overexpressing (Ankrd2(+)clS9 and Ankrd2(+)clS12) and two –silencing clones (Ankrd2(-)clA9 and Ankrd2(-)clA10)and from C2C12 control cells at proliferating myoblasts stage. Competitive hybridization was done twice using different microarray slides in which each single sample and control RNA was labeled either with Cy3 or Cy5 fluorochromes (dye swapping procedure).Using this strategy we obtained four fluorescence intensity values for each probe in the platform. Fluorescence detection was carried out on a GSI Lumonics LITE dual confocal laser scanner (Perkin-Elmer) with ScanArray Microarray Analysis Software, and raw scanner images were analyzed with QuantArray software (Perkin-Elmer). Normalization of the expression levels was performed with MIDAW tool (Romualdi et al., 2005).
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| Contributor(s) |
Bean C, Facchinello N, Salamon M, Faulkner G, Lanfranchi G |
| Citation(s) |
18302940 |
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| Submission date |
Apr 18, 2007 |
| Last update date |
Mar 16, 2012 |
| Contact name |
Gerolamo Lanfranchi |
| E-mail(s) |
stefano.cagnin@unipd.it
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| Phone |
+39-0498276219
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| Organization name |
University of Padova
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| Department |
CRIBI - Biotechnology Center and Biology Department
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| Lab |
Functional Genomics Lab
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| Street address |
Via U. Bassi, 58/B
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| City |
Padova |
| ZIP/Postal code |
35131 |
| Country |
Italy |
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| Platforms (1) |
| GPL5098 |
Micro-CRIBI Mouse 29K Oligo Array (Operon V1.1) |
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| Samples (8)
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| GSM182653 |
Gene expression profiling of Ankrd2(-)clA10 (A) |
| GSM182748 |
Gene expression profiling of Ankrd2(-)clA10 (B) |
| GSM182749 |
Gene expression profiling of Ankrd2(-)clA9 (A) |
| GSM182750 |
Gene expression profiling of Ankrd2(-)clA9 (B) |
| GSM182751 |
Gene expression profiling of Ankrd2(+)clS12 (A) |
| GSM182752 |
Gene expression profiling of Ankrd2(+)clS12 (B) |
| GSM182753 |
Gene expression profiling of Ankrd2(+)clS9 (A) |
| GSM182754 |
Gene expression profiling of Ankrd2(+)clS9(B) |
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| Relations |
| BioProject |
PRJNA99373 |
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