Tβ-4 CRISPR Activation Plasmid (h): sc-417057-ACT

Thymosin beta-4, encoded by human TMSB4X, is a highly conserved actin-sequestering peptide that regulates G-actin availability and supports cytoskeletal remodeling, cell motility, and stress-responsive changes in cell shape. By modulating actin dynamics, it influences processes such as migration, adhesion, and wound-associated cellular responses, and has been linked to pathways controlling inflammation and tissue remodeling. Altered TMSB4X expression has been reported across contexts including tumor progression, angiogenic signaling, and fibrotic or inflammatory states, making it a useful node for mechanistic studies of cytoskeleton-dependent phenotypes. As a broadly expressed cytoskeletal regulator, TMSB4X provides a tractable target for interrogating how actin homeostasis interfaces with transcriptional programs and cellular microenvironment cues.

Tβ-4 CRISPR Activation Plasmid (h) provides a targeted, non-destructive approach to upregulating endogenous TMSB4X expression without altering the underlying DNA sequence.

Tβ-4 CRISPR Activation Plasmid (h) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the TMSB4X 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 TMSB4X transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous Tβ-4 expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native TMSB4X locus and enabling the study of Tβ-4-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of Tβ-4 pathway restoration in tumor cells with silenced or reduced TMSB4X expression.

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