Jujube (Ziziphus jujuba Mill.) is a traditional fruit tree in China with immense economic and ecological value. Jujube fruits are abundant in polyphenolic secondary metabolites, particularly proanthocyanidins (PAs), which play a crucial role in enhancing the quality of jujube fruits. However, the mechanism underlying the biosynthesis of PAs remains unclear. The PA contents of sour jujube ‘Qingjiansuanzao’ and cultivated jujube ‘Junzao’ were compared at different developmental stages to unravel this mechanism. The PA contents of sour jujube were higher than that of cultivated jujube and decreased during fruit development. Combined with transcriptome analysis, a large number of differentially expressed genes related to PA biosynthesis were screened. Correlation analysis showed that ZjLAR and ZjANR played an active role in promoting the biosynthesis of PAs. Transient overexpression of ZjLAR and ZjANR in jujube fruits resulted in higher total PAs and monomeric catechin, but the PAs decreased after transient silencing. Overexpressing ZjLAR and ZjANR in Arabidopsis and tomato increased the content of PAs in Arabidopsis seeds and tomato fruits. These findings provide a new basis for further understanding of the biosynthesis of jujube PAs and are significant for improving the quality of jujube fruit.