Transcriptome analysis of green and purple fruited pepper provides insight into novel regulatory genes in anthocyanin biosynthesis

Background: Pepper (Capsicum annuum L.) is a valuable horticultural crop with economic significance, and its purple fruit color is attributed to anthocyanin, a phytonutrient known for its health-promoting benefits. However, the mechanisms regulating anthocyanin biosynthesis in pepper have yet to be fully elucidated.

Methods: RNA sequencing (RNA-seq) was utilized to analyze the transcriptome of fruits from three purple-fruited varieties (HN191, HN192, and HN005) and one green-fruited variety (EJT) at various developmental stages. To determine the relationships between samples, Pearson correlation coefficients (PCC) and principal component analysis (PCA) were calculated. Differential expression analysis was performed using the DESeq2 package to identify genes that were expressed differently between two samples. Transcription factors (TF) were predicted using the iTAK program. Heatmaps of selected genes were generated using Tbtools software.

Results: The unripe fruits of HN191, HN192, and HN005, at the stages of 10, 20, and 30 days after anthesis (DAA), display a purple color, whereas the unripe fruits of variety EJT remain green. To understand the molecular basis of this color difference, five transcriptome comparisons between green and purple fruits were conducted: HN191-10 vs EJT-10, HN191-20 vs EJT-20, HN191-30 vs EJT-30, HN192-30 vs EJT-30, and HN005-30 vs EJT-30. Through this analysis, 503 common differentially expressed genes (DEGs) were identified. Among these DEGs, eight structural genes related to the anthocyanin biosynthesis pathway and 24 transcription factors (TFs) were detected. Notably, one structural gene (MSTRG.12525) and three TFs (T459_25295, T459_06113, T459_26036) exhibited expression patterns that suggest they may be novel candidate genes involved in anthocyanin biosynthesis. These results provide new insights into the regulation of anthocyanin biosynthesis in purple pepper fruit and suggest potential candidate genes for future genetic improvement of pepper germplasm with enhanced anthocyanin accumulation.