Stabilization of cyclin D3 protein by CDKN1A (p21

Butyrate-mediated inhibition of cell proliferation is part of the preventive role of dietary fiber against colorectal cancer (CRC). This effect notably involves the cyclin-dependent kinase inhibitor CDKN1A (p21Cip/Waf1) in human intestinal cells, yet the underlying molecular mechanisms remain incompletely understood. Previously, we observed a paradoxical increase in cyclin D3 (CCND3)—but not cyclin D1—levels upon butyrate exposure. Here, we demonstrate that the butyrate-induced accumulation of CCND3 protein results both from mRNA increase and a CDKN1A-dependent protein stabilization, specifically extending its nuclear half-life. Proteomic analyses of CCND3 co-immunoprecipitates identified complexes particularly enriched in butyrate-treated cells and involving CDKN1A, CDK4, CDK6, and the CRC-associated kinase CDK5. Phosphorylation at a conserved Thr residue in the C-terminal intrinsically disordered region, crucial for CCND nuclear export and subsequent proteasomal degradation, was notably reduced following butyrate treatment and inversely correlated with CDKN1A expression levels. Structural modeling based on AlphaFold2, complemented by molecular dynamics simulations, revealed possible differential interactions between CDKN1A and cyclins D1 and D3, predicting that CCND3-Thr283 becomes structurally buried upon CDKN1A binding, limiting its phosphorylation. Our findings suggest novel mechanistic insights into how the droplet driver CDKN1A might regulate CCND3 stability, highlighting previously unexplored roles of cyclin D3-containing complexes in cell cycle arrest induced by butyrate.