An integrin a3ß1-CSTF3 signaling axis regulates alternative polyadenylation of Mmp9 mRNA

The laminin-binding integrin a3ß1 is highly expressed in epidermal keratinocytes, where it coordinates diverse cellular functions and gene expression during skin remodeling. Here, we show that a3ß1-MEK/ERK signaling operates in vivo to promote proximal polyadenylation site (PAS) usage in the Mmp9 gene, generating a short, more stable mRNA transcript. Using mice with inducible, epidermis-specific a3 deletion, RNA in situ hybridization revealed that loss of a3ß1 increased the long Mmp9 transcript in healing wounds and epidermal tumors. a3ß1-MEK/ERK signaling in keratinocytes induced the expression of the cleavage stimulation factor CSTF3, a known regulator of alternative polyadenylation (APA), while CSTF3 knockdown shifted Mmp9 toward distal PAS usage. Moreover, a3 deletion reduced Cstf3 gene expression and altered APA in vivo. Genome-wide DaPars2 analysis identified a3ß1-dependent APA across numerous genes, including some encoding components of the keratinocyte secretome. Together, these findings define a novel a3ß1-MEK/ERK-CSTF3 axis that orchestrates post-transcriptional gene regulation through APA, revealing a3ß1 as a potential target for wound and cancer therapies. Overall design: Immortalized mouse keratinocytes (MK cells) that lack a3 integrin (MK-/-), and MK-/- cells reconstituted with human a3 integrin (MK-/-:ha3), were cultured on collagen-coated tissue culture dishes in supplemented low calcium Minimum Essential Medium (MEM; Thermo Fisher Scientific; cat. # 11380037) and incubated at 33oC with 8% CO2. RNA isolation and sequencing was performed by Azenta Life Sciences. All downstream analyses were conducted by the Albany Medical College Bioinformatics Core, including quality control (FastQC, MultiQC), read alignment, gene quantification, and integrative analyses.