| RRC ID |
30717
|
| 著者 |
Yoshimoto N, Tatematsu K, Iijima M, Niimi T, Maturana AD, Fujii I, Kondo A, Tanizawa K, Kuroda S.
|
| タイトル |
High-throughput de novo screening of receptor agonists with an automated single-cell analysis and isolation system.
|
| ジャーナル |
Sci Rep
|
| Abstract |
Reconstitution of signaling pathways involving single mammalian transmembrane receptors has not been accomplished in yeast cells. In this study, intact EGF receptor (EGFR) and a cell wall-anchored form of EGF were co-expressed on the yeast cell surface, which led to autophosphorylation of the EGFR in an EGF-dependent autocrine manner. After changing from EGF to a conformationally constrained peptide library, cells were fluorescently labeled with an anti-phospho-EGFR antibody. Each cell was subjected to an automated single-cell analysis and isolation system that analyzed the fluorescent intensity of each cell and automatically retrieved each cell with the highest fluorescence. In ~3.2 × 10(6) peptide library, we isolated six novel peptides with agonistic activity of the EGFR in human squamous carcinoma A431 cells. The combination of yeast cells expressing mammalian receptors, a cell wall-anchored peptide library, and an automated single-cell analysis and isolation system might facilitate a rational approach for de novo drug screening.
|
| 巻・号 |
4
|
| ページ |
4242
|
| 公開日 |
2014-2-28
|
| DOI |
10.1038/srep04242
|
| PII |
srep04242
|
| 解説 |
S.cerevisiaeでGal1プロモーター制御下での目的遺伝子の発現に使用できます。本研究ではアゴニスト探索のためにEGFRを酵母で発現するために使用されました。
|
| PMID |
24577528
|
| PMC |
PMC3937795
|
| MeSH |
Biological Assay / instrumentation*
Biological Assay / methods
Cell Separation / instrumentation*
Cell Separation / methods
Drug Evaluation, Preclinical / instrumentation*
Drug Evaluation, Preclinical / methods
Equipment Design
Equipment Failure Analysis
Flow Cytometry / instrumentation*
Flow Cytometry / methods
High-Throughput Screening Assays / instrumentation*
High-Throughput Screening Assays / methods
Receptors, Cytoplasmic and Nuclear / antagonists & inhibitors*
Robotics / instrumentation*
Robotics / methods
|
| IF |
3.998
|
| 引用数 |
6
|
|
WOS 分野
|
BIOCHEMISTRY & MOLECULAR BIOLOGY
|
| リソース情報 |
| 遺伝子材料 |
pGMH20 (RDB01956). |
