Trichohyalin CRISPR Activation Plasmid (h): sc-401940-ACT

TCHH encodes trichohyalin, a highly crosslinked structural protein enriched in the inner root sheath of hair follicles and in differentiating keratinocytes. Trichohyalin is a major substrate for peptidylarginine deiminases and transglutaminases, supporting citrullination and isopeptide crosslinking that stabilize the cornified envelope and reinforce epithelial barrier mechanics. Through its integration with keratin intermediate filaments and the keratinization program, TCHH contributes to follicular morphogenesis, hair shaft architecture, and epidermal differentiation. Dysregulated expression or processing of trichohyalin has been associated with hair and skin phenotypes, making it relevant for studies of follicle biology, barrier disorders, and epithelial stress responses.

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

Trichohyalin CRISPR Activation Plasmid (h) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the TCHH 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 TCHH transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous Trichohyalin expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native TCHH locus and enabling the study of Trichohyalin-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of Trichohyalin pathway restoration in tumor cells with silenced or reduced TCHH expression.

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