Genes and compounds that increase type VII collagen expression as potential treatments for dystrophic epidermolysis...

Area of Science:

• Genetics
• Dermatology
• Molecular Biology

Background:

• Dystrophic Epidermolysis Bullosa (DEB) is a severe genetic skin disorder caused by mutations in the COL7A1 gene, leading to deficient type VII collagen (C7).
• Current treatments for DEB are limited, and there is no cure; however, increasing functional C7 production shows therapeutic promise.
• CRISPR-Cas9 technology has proven effective in identifying genes influencing various cellular processes, including disease mechanisms.

Purpose of the Study:

• To identify genes and pathways that can enhance the expression of type VII collagen (C7) in keratinocytes.
• To discover small molecules that upregulate C7 production for potential DEB therapies.
• To validate a reporter cell line for efficient screening of C7 expression modifiers.

Main Methods:

• Development of a keratinocyte cell line engineered to report C7 expression levels (C7_tdTomato).
• Genome-wide CRISPR activation (CRISPRa) screen to identify genetic modifiers of C7 expression.
• Targeted drug screen using identified pathways and compounds to find C7-upregulating agents.

Main Results:

• The CRISPRa screen identified DENND4B and TYROBP as key genes involved in C7 regulation, alongside pathways related to calcium uptake and immune signaling.
• A targeted drug screen revealed several compounds capable of increasing C7 expression in keratinocytes.
• The plant flavonoid kaempferol demonstrated significant upregulation of C7 mRNA and protein in DEB patient-derived cells.

Conclusions:

• CRISPRa screening is a viable method for discovering regulators of C7 production.
• DENND4B, TYROBP, calcium signaling, and immune pathways are implicated in controlling C7 levels.
• Kaempferol represents a promising therapeutic candidate for Dystrophic Epidermolysis Bullosa due to its ability to restore C7 expression in patient cells.