Tomato ( Solanum lycopersicum) is a popular vegetable crop worldwide, valued for its high nutritional content and significant economic importance. However, low-temperature stress is a major problem for growing tomato in cooler regions, as it reduces yield and sometimes plant death, resulting in considerable losses in production. Therefore, exploring the genes mediating cold tolerance and deploying them in breeding cold-tolerant tomato varieties are essential for sustaining and expanding tomato production. Basic transcription factor 3 (BTF3), the β-subunit (βNAC) of the nascent-polypeptide-associated complex, has been linked to stress tolerance in other species, but its role in tomato cold adaptation is unknown. To elucidate the role of BTF3 in cold tolerance in tomato, we generated SlBTF3-overexpression ( OE) lines and generated slbtf3 knock-out mutants. It was found that SlBTF3-OE plants were highly cold-tolerant, while slbtf3 mutant exhibited greater susceptibility to low temperatures compared to wild-type. Mass spectrometry analysis revealed that SlBTF3 interacts with SlCBF1, a key component in cold stress responses, and stabilizes its expression, while the protein kinase SlOST1 interacts with and phosphorylates SlBTF3. Further analysis revealed that elevated SlBTF3 expression preserves chloroplast integrity under cold stress by upregulating and stabilizing SlCBF, which in turn enhances the activity of antioxidant enzymes. Furthermore, SlBTF3-OE plants exhibited higher expression levels of PsaA, PsaB, PsaC, and PetB, key components of photosystem I and the cytochrome b 6f complex. In summary, this study enhances our understanding of SlBTF3-mediated cold tolerance in tomato, laying the groundwork for breeding stress-resilient tomato cultivars.