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| Status |
Public on Jan 10, 2018 |
| Title |
A method for the acute and rapid degradation of endogenous protein |
| Organism |
Mus musculus |
| Experiment type |
Expression profiling by high throughput sequencing
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| Summary |
We developed Trim-Away as a a technique to degrade endogenous proteins acutely in mammalian cells without prior modification of the genome or mRNA. Trim-Away is able to remove unmodified native proteins within minutes of application by making use of the TRIM21 ubiquitin ligase that regognises antibody-protein complexes and harnesses the cellular degradation machinery. We performed transcriptome profiling (RNAseq) on NIH3T3 and NIH3T3-mCherry-mTRIM21 cells following electroporation with non-targeting antibody (IgG), bovine serum albumin (BSA), or buffer (PBS). We found that a total of only 8 protein-encoding transcripts were consistently downregulated more than 2-fold in the NIH3T3-mCherry-mTRIM21 cells (Kdm5d, Ddx3y, Eif2s3y, Uty, Asb4, Fat4, Papss2 and Lama2), although we cannot rule out the possibility that this is an indirect consequence of lentivirus construct integration rather than a direct consequence of TRIM21 overexpression. Notably, transcripts encoding previously described ligands of TRIM21, IRF-3, IRF-5, Skp2, DAXX, DDX41, and SQSTM1 were not significantly increased or decreased upon overexpression of TRIM21. Our experiments suggest that the cells overexpressing TRIM21 behave similar to wild-types.
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| Overall design |
mRNA profiles of NIH3T3 and NIH3T3-mCherry-mTRIM21 cells following electroporation with antibody (IgG), BSA or PBS were generated by deep sequencing, in triplicate, using a Illumina NextSeq 500 platform.
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| Contributor(s) |
Clift D, McEwan WA, Labzin LI, Konieczny V, Mogessie B, James LC, Schuh M |
| Citation(s) |
29153837 |
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| Submission date |
Oct 20, 2017 |
| Last update date |
Jul 25, 2021 |
| Contact name |
Melina Schuh |
| Organization name |
Max Planck Institute for Biophysical Chemistry
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| Street address |
Am Fassberg 11
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| City |
Göttingen |
| ZIP/Postal code |
37077 |
| Country |
Germany |
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| Platforms (1) |
| GPL19057 |
Illumina NextSeq 500 (Mus musculus) |
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| Samples (18)
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| Relations |
| BioProject |
PRJNA415216 |
| SRA |
SRP120628 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSE105471_RAW.tar |
1.7 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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