Reporter Cell Lines
Membrane protein stable cell lines are widely used in many areas of biomedical research. Creative Biolabs can offer membrane protein stable cell lines to stablish in vitro models for High Throughput Screening.
Creative Biolabs offers high-quality, innovative tools to help research groups accelerate membrane protein drug discovery. They can be found by targets. If there is no product that meets your needs, please contact us.
Background of Reporter
The gene promoter regions contain responsive elements for second messengers that modify gene transcription when GPCRs are activated, including the cAMP response element (CRE), the nuclear factor of activated T-cells response element (NFAT-RE), the serum response element (SRE), and the serum response factor response element (SRF-RE, a mutant form of SRE). Cell-based reporter assays thus provide a different popular and cost-effective HTS platform for GPCR screening. Enzyme proteins like luciferase, alkaline phosphatase, -galactosidase, and -lactamase, as well as different fluorescent proteins, are frequently utilized as reporters because their activities can be connected to a variety of colorimetric, fluorescent, or luminous readouts.
Advantages of Reporter Cell Lines
The wide linearity and sensitivity of reporter gene assays, as well as their high signal-to-background ratio, make them ideal for the detection of weak GPCR agonists or allosteric modulators. In addition to being simple to set up, reporter gene assays can be scaled down to very small test volumes in 1536- or 3456-well formats. Despite these benefits, numerous issues have been brought up, including the need for lengthy incubation times, the challenge of antagonist detection due to reporter accumulation, and the increased risk of false positives because the signal event occurs later than receptor activation. By employing destabilized reporters, issues with the protracted incubation period and accumulation of reporter have been addressed. By co-expressing an internal control that is constitutively expressed, the greater false positive rate brought on by the distal signaling event could be partially mitigated, allowing for the exclusion of chemicals that alter gene transcription in an unintended manner.
Published Data
| Paper Title | TRACE generates fluorescent human reporter cell lines to characterize epigenetic pathways |
| Journal | Molecular Cell |
| Published | 2022 |
| Abstract | Although genetically encoded biosensors are effective tools for tracking cellular activity, the ability to understand epigenetic pathways is hampered by the challenge of producing suitable reporters for chromatin factors. In this article, researchers present TRACE (transgene reporters across chromatin environments), a high-throughput, genome-wide method for creating fluorescent human reporter cell lines responsive to altering epigenetic factors. They identify reporters responsive to pharmacological inhibition of the histone lysine demethylase LSD1 and genetic ablation of the PRC2 subunit SUZ12 by profiling GFP expression from a large pool of individually barcoded lentiviral integrants in the presence and absence of a perturbation. Additionally, by modifying the HIV-1 host factor LEDGF through targeted deletion or fusion to chromatin reader domains, they change the preferences for lentiviral integration sites, expanding the range of chromatin types that TRACE can analyze. Their mechanistic understanding of chromatin biology will advance thanks to the genetic interrogation of a wide variety of epigenetic pathways made possible by the phenotypic reporters created by TRACE. |
| Result |
To identify reporters sensitive to pharmacological disruption of LSD1-mediated histone lysine demethylation, they carried out a proof-of-concept TRACE experiment. The removal of mono- and di-methyl groups from histone H3's lysine 4 is catalyzed by LSD1 (H3K4me1 and H3K4me2). Thus, the LSD1 inhibitor GSK2879552 (also known as LSD1i) causes transcriptional derepression and an accumulation of H3K4me1 and H3K4me2 at LSD1 target sites in treated cells.
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Fig.1 TRACE identifies reporters responsive to LSD1 inhibition.1,2
References
- Tchasovnikarova, Iva A., et al. "TRACE generates fluorescent human reporter cell lines to characterize epigenetic pathways." Molecular cell 82.2 (2022): 479-491.
- Distributed under Open Access License CC BY 4.0, without modification.
