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Part:BBa_K5267048

Designed by: Zhang Renjie   Group: iGEM24_NUDT-CHINA   (2024-09-29)
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P_CMV->MTNR1B->bGH_polyA

Expression of MT2 gene

Sequence and Features


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    COMPATIBLE WITH RFC[12]
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal BglII site found at 614
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    COMPATIBLE WITH RFC[25]
  • 1000
    COMPATIBLE WITH RFC[1000]


Profile

Name: P_CMV->MTNR1B->bGH_polyA
Base Pairs: 1949bp
Origin: Homo sapiens
Properties: Expression of MT2 gene

Usage and Biology

The melatonin receptors (MTs), specifically MT1 (melatonin receptor type 1) and MT2 (melatonin receptor type 2), are classified under the G protein-coupled receptor (GPCR) family A, with melatonin acting as their endogenous agonist. These receptors are pivotal in the regulation of the circadian rhythm within the human body and are intricately linked to a spectrum of vital physiological processes, including reproductive function, neuronal modulation, and immune system regulation. Furthermore, MTs represent a significant therapeutic target for the amelioration of various pathologies, such as insomnia, affective disorders, and oncological conditions.[1].

Despite the high degree of homology between human MT1 and MT2 receptors, there are considerable differences in their tissue distribution, intracellular signaling mechanisms, and physiological roles. The current dearth of selectivity in melatonin-based pharmaceuticals for either MT1 or MT2 receptors impedes the precision therapy of related disorders. Consequently, the detailed structural elucidation of the MTs agonist binding site is of paramount importance for the development of targeted pharmacotherapeutics.[2]

In light of this, we have engineered a pathway that, upon promoter activation, initiates the synthesis of the MT2 melatonin receptor protein. This strategy is instrumental in the establishment of a cellular assay system designed for the screening of melatonin receptor agonists.(Figure 1)


Figure 1. MT2 gene expression pathway.

Special design

To achieve the objective of driving the expression of the MT2 melatonin receptor, we strategically selected the CMV promoter, a robust promoter derived from the human Cytomegalovirus (CMV), known for its high transcriptional activity in eukaryotic cells. The CMV promoter has been demonstrated to be highly efficacious in facilitating the expression of lengthy and complex genes within HEK-293T cells.[3]

Utilizing the CMV promoter, we initiated the transcription of the MT2 gene within the construct of the gene expression vector, thereby enhancing the expression profile of the MT2 gene. This approach is anticipated to provide a foundation for the development of a cell-based screening platform for melatonin receptor agonists.

Reference

[1] “Melatonin receptor structure and signaling,” J. Pineal Res., vol. 76, no. 3, p. e12952, Apr. 2024, doi: 10.1111/jpi.12952.
[2] S. Tordjman et al., “Melatonin: Pharmacology, Functions and Therapeutic Benefits,” Curr. Neuropharmacol., vol. 15, no. 3, pp. 434–443, Feb. 2017, doi: 10.2174/1570159X14666161228122115.
[3] A. H. Rad S. M., A. Poudel, G. M. Y. Tan, and A. D. McLellan, “Promoter choice: Who should drive the CAR in T cells?,” PLOS ONE, vol. 15, no. 7, p. e0232915, Jul. 2020, doi: 10.1371/journal.pone.0232915.

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