TSHR CRISPR Activation Plasmid (h): sc-400848-ACT

Thyroid stimulating hormone receptor (TSHR) is a glycosylated class A GPCR expressed primarily in thyroid follicular cells, where it transduces pituitary TSH signals to regulate thyroid hormone biosynthesis and gland growth. Upon ligand engagement, TSHR activates Gαs-driven cAMP/PKA signaling and can also engage Gαq/PLC and MAPK pathways, coordinating transcriptional programs controlling iodide uptake, thyroglobulin processing, and thyroid-specific differentiation. Dysregulated TSHR signaling or expression is linked to autoimmune thyroid disease, including Graves’ disease, and to altered thyroid homeostasis that can contribute to hyperfunctioning nodules and neoplastic transformation. As a surface receptor integrating endocrine and immune-related cues, TSHR is a key model for studying GPCR trafficking, receptor desensitization, and thyroid lineage regulatory networks.

TSHR CRISPR Activation Plasmid (h) provides a targeted, non-destructive approach to upregulating endogenous TSHR expression without altering the underlying DNA sequence.

TSHR CRISPR Activation Plasmid (h) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the TSHR locus in human cell lines. The system is built around a catalytically inactive Cas9 (dCas9) carrying two inactivating mutations (D10A and N863A) that eliminate nuclease activity while preserving DNA binding. This dCas9 is fused to VP64, a potent transcriptional activator, and is co-expressed with a blasticidin resistance gene for selection. The second plasmid encodes the MS2-p65-HSF1 fusion protein, a secondary activator complex that works in concert with dCas9-VP64, alongside a hygromycin resistance gene. The third plasmid encodes a target-specific 20 nt sgRNA fused to two MS2 RNA aptamers that recruit the MS2-p65-HSF1 complex to the activation site, accompanied by a puromycin resistance gene. The three plasmids are delivered at a 1:1:1 mass ratio for balanced expression of all system components.

Once assembled at the target locus, the SAM complex binds within approximately 200 bp upstream of the TSHR transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous TSHR expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native TSHR locus and enabling the study of TSHR-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of TSHR pathway restoration in tumor cells with silenced or reduced TSHR expression.

For Research Use Only. Not Intended for Diagnostic or Therapeutic Use.