Reprogramming signal transduction through a designer receptor tyrosine kinase

Kongkrongtong, Tatphon; Sumigama, Yuka; Nagamune, Teruyuki; Kawahara, Masahiro

doi:10.1038/s42003-021-02287-8

Cited by 6 publications

(5 citation statements)

References 29 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…2 c). The result is consistent with our recent study showing that STAT3 or STAT5, but not STAT1, promotes proliferation of Ba/F3 cells 25 , 26 . These results indicate that the engineered receptor scaffold activates signaling molecules bound to the tyrosine motif in response to the synthetic ligand AP20187.…”

Section: Resultssupporting

confidence: 94%

“…The signaling analyses of the selected tyrosine motif clones demonstrated that proliferation-inducing signaling molecules such as STAT5, MEK, and Akt were activated in both Ba/F3 and 32Dcl3 cells. As shown in our previous study, proliferation of Ba/F3 cells is promoted by STAT3 or STAT5 25 , 26 . The introductory experiments (Fig.…”

Section: Discussionsupporting

confidence: 66%

“…1 ). A backbone plasmid pMK-stuffer-IPTG 25 was used for retroviral infection, in which internal ribosomal entry site (IRES; I) is used for co-expression of the genes encoding a puromycin (P) resistance, a self-cleaving T2A peptide (T), and an enhanced green fluorescence protein (EGFP; G). The open reading frame encoding each receptor sequence was amplified with PCR and inserted into pMK-stuffer-IPTG using In-fusion HD Cloning Kit (TakaraBio).…”

Section: Methodsmentioning

confidence: 99%

See 2 more Smart Citations

Phenotypic screening of signaling motifs that efficiently induce cell proliferation

Umene,

Nagamune,

Kawahara

2023

Sci Rep

Self Cite

View full text Add to dashboard Cite

Since cell proliferation is one of the fundamental cell fates, artificial control of cell proliferation based on a receptor-engineering approach is increasingly important in therapeutic and industrial applications. Since the signal transduction properties of cytokine receptors are greatly influenced by the amino acid sequence of tyrosine motifs, here we develop a phenotypic screening approach that can directly select cell proliferation-inducing tyrosine motifs from a synthetic library. In the tyrosine motif library, amino acid sequences around the tyrosine are randomized to attain diverse binding patterns of signaling molecules. Theoretically, engineered receptors with distinct tyrosine motifs would activate signaling molecules in diverse patterns. Thus, we investigated whether tyrosine motif sequences capable of inducing cell proliferation could be selected from the cellular library expressing the motif-engineered receptors. Consequently, the selected motifs induced similar levels of cell proliferation compared to the cytoplasmic signaling domain of a native receptor. The motif-screening system was applicable to cells that may differentiate or proliferate depending on cytokine signals. To our best knowledge, this is the first report demonstrating phenotypic screening of tyrosine motifs in living cells. Our approach would open up new possibilities in the field of artificial control of cell fate based on signal transduction engineering.

show abstract

Section: Resultssupporting

confidence: 94%

Section: Discussionsupporting

confidence: 66%

Section: Methodsmentioning

confidence: 99%

See 1 more Smart Citation

Phenotypic screening of signaling motifs that efficiently induce cell proliferation

Umene,

Nagamune,

Kawahara

2023

Sci Rep

Self Cite

View full text Add to dashboard Cite

show abstract

“…However, we never know how much levels the signaling molecules should be phosphorylated for cell proliferation. Our previous studies using chimeric receptors demonstrated that Ba/F3, which is a host cell line of THROPPIS, proliferated when either MEK, STAT3, or STAT5 was activated Kongkrongtong et al, 2021). In addition, the chimeric receptors in THROPPIS are not embedded in the membrane but expressed in the cytosol, which means that the signaling pathways could be different from those of the natural receptor TPOR.…”

Section: Throppis Can Be Extended To Other Receptorsmentioning

confidence: 99%

Thrombopoietin receptor‐based protein–protein interaction screening (THROPPIS)

Horikawa

Kakiuchi

Kashima

et al. 2021

Biotech & Bioengineering

Self Cite

View full text Add to dashboard Cite

As protein-protein interactions (PPIs) are involved in many cellular events, development of mammalian cytosolic PPI detection systems is important for drug discovery as well as understanding biological phenomena. We have previously reported a c-kitbased PPI screening (KIPPIS) system, in which proteins of interest were fused with a receptor tyrosine kinase c-kit, leading to intracellular PPI-dependent cell growth.However, it has not been investigated whether PPI can be detected using other receptors. In this study, we employed a thrombopoietin receptor, which belongs to the Type I cytokine receptor family, to develop a thrombopoietin receptor-based PPI screening (THROPPIS) system. To improve the sensitivity of THROPPIS, we examined two strategies of (i) localization of the chimeric receptors on the cell membrane, and (ii) addition of a helper module to the chimeric receptors. Intriguingly, the nonlocalized chimeric receptor showed the best performance of THROPPIS. Furthermore, the addition of the helper module dramatically improved the detection sensitivity. In total, 5 peptide-domain interactions were detected successfully, demonstrating the versatility of THROPPIS. In addition, a peptide-domain interaction was detected even when insulin receptor or epidermal growth factor receptor was used as a signaling domain, demonstrating that this PPI detection system can be extended to other receptors.

show abstract

“…Particularly, iCasp9, an inducible suicide gene, has progressed to Phase I clinical trials as a safety switch in cell therapy 17 . In addition, we previously developed designer receptors that can activate target signaling molecules by appropriately arranging domains and tyrosine motifs 18 – 20 . Such a designer receptor was also utilized for phenotypic screening of tyrosine motifs that efficiently induce cell proliferation 21 .…”

Section: Introductionmentioning

confidence: 99%

Synthetic receptor scaffolds significantly affect the efficiency of cell fate signals

Umene,

Kawahara

2024

Sci Rep

Self Cite

View full text Add to dashboard Cite

Mimicry of receptor functions by designing synthetic receptors would be one of the recently hot research trends in cell engineering. While several types of synthetic receptors have been designed to induce desired cell fates in response to external stimuli, little is known about which receptor type signals more efficiently for inducing a certain cell fate. In this study, we compared the performance of three types of synthetic receptor scaffolds, i.e. myristoylated, cytosolic, and transmembrane types that signal through JAK-dependent phosphorylation of tyrosine motifs to transduce growth signaling. As a result, the phosphorylation levels of JAK and subsequent downstream signaling molecules were significantly maintained in the cytosolic type receptors, leading to more efficient cell growth than the other types. In contrast, the phosphorylation levels of JAK decreased in a motif-dependent manner in the transmembrane type receptors. Although various studies on receptor engineering based on domain or motif engineering have been reported, to our knowledge this study is the first to demonstrate that synthetic receptor scaffolds significantly affect the efficiency of cell fate signals. These findings are important for both receptor biology and receptor engineering, providing guidelines for rationally designing synthetic receptors that can transduce as efficient signaling as possible.

show abstract

Reprogramming signal transduction through a designer receptor tyrosine kinase

Cited by 6 publications

References 29 publications

Phenotypic screening of signaling motifs that efficiently induce cell proliferation

Phenotypic screening of signaling motifs that efficiently induce cell proliferation

Thrombopoietin receptor‐based protein–protein interaction screening (THROPPIS)

Synthetic receptor scaffolds significantly affect the efficiency of cell fate signals

Contact Info

Product

Resources

About