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| Status |
Public on Feb 01, 2016 |
| Title |
A Dual Molecular Analog Tuner for Dissecting Mammalian Protein Function |
| Organism |
Homo sapiens |
| Experiment type |
Expression profiling by high throughput sequencing
|
| Summary |
Loss-of-function studies are fundamental for dissecting gene function. Yet, methods to rapidly and effectively perturb genes in mammalian cells are scarce. We present a novel system, deliverable with only two lentiviral vectors, which enables simultaneous control over two different proteins in the same cell. By harnessing the plant auxin and jasmonate hormone-induced degradation pathways, combined with RNA interference, this system allows constitutive depletion of two endogenous proteins and their replacement with two exogenous proteins whose degradation is rapidly and reversibly induced by external ligands, representing a dual analog molecular tuner. Focusing on NANOG, CHK1 and NOTCH1 in embryonic stem cells and p53 in cancer cells we have validated the efficiency, rapidity, reversibility, titratability and multiplicity of the engineered tuners, and demonstrated their potential to facilitate previously-unfeasible experimental approaches and to generate novel biological insights.
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| |
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| Overall design |
For mRNA-Seq preparation, coronatine/DMSO treated cells were collected.
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| |
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| Contributor(s) |
Brosh R, Waghray A, Lemischka IR |
| Citation(s) |
27230261 |
| NIH grant(s) |
| Grant ID |
Grant title |
Affiliation |
Name |
| R01 GM078465 |
Global & Systems Level Analyses of Cell Fate Regulation |
MOUNT SINAI SCHOOL OF MEDICINE |
IHOR R LEMISCHKA |
|
| Submission date |
Oct 28, 2015 |
| Last update date |
May 15, 2019 |
| Contact name |
avinash waghray |
| E-mail(s) |
avinash.waghray@gmail.com
|
| Organization name |
Massachussets General Hospital
|
| Department |
Center for Regenerative Medicine
|
| Lab |
Rajagopal
|
| Street address |
185 Cambridge Street, CPZN Room 4254A
|
| City |
Boston |
| State/province |
MA |
| ZIP/Postal code |
02114 |
| Country |
USA |
| |
|
| Platforms (1) |
| GPL18573 |
Illumina NextSeq 500 (Homo sapiens) |
|
| Samples (4)
|
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| Relations |
| BioProject |
PRJNA300451 |
| SRA |
SRP065451 |
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