Mlx CRISPR Activation Plasmid (h): sc-403499-ACT

MLX encodes Mlx, a basic helix–loop–helix leucine zipper transcription factor that forms obligate heterodimers with Mondo family partners such as MLXIP (MondoA) and MLXIPL (ChREBP) to couple nutrient availability to gene expression. These complexes regulate carbohydrate and lipid metabolism programs, including glucose-responsive transcription and glycolytic/lipogenic gene networks, linking mitochondrial and cytosolic metabolite sensing to transcriptional control. Through interactions with broader MYC/MAX/MLX transcriptional circuitry, Mlx contributes to coordination of growth, energy homeostasis, and stress-adaptive responses. Dysregulation of MLX-associated transcriptional programs has been implicated in metabolic disease–relevant pathways and tumor cell metabolic rewiring, supporting its use in mechanistic studies of transcriptional control in metabolism and proliferative states.

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

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

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