Fumonisins, a major class of mycotoxins, pose significant health risks to humans and animals due to their widespread contamination and potent toxicity. Recent advances in molecular biology, biochemistry, and enzymology have greatly enhanced the understanding of fumonisin biosynthesis and its genetic regulation. The key biosynthetic genes are typically organized in clusters and regulated by specific transcription factors; increasing evidence also highlights the involvement of complex transcriptional and epigenetic mechanisms. Environmental factors such as nitrogen, carbon, and pH also modulate these regulatory networks. Despite substantial progress, critical gaps remain in fully elucidating the regulatory pathways that control fumonisin production. This review synthesizes current knowledge regarding fumonisin biosynthesis, gene clusters, and multi-level regulatory mechanisms, while emphasizing recent trends, existing challenges, and potential applications in food safety and biotechnology to enhance food security and promote sustainable development.