GhCTSF1, a short PPR protein with a conserved role in chloroplast development and photosynthesis, participates in intron splicing of rpoC1 and ycf3-2 transcripts in cotton

Cotton is one of the most important textile fibers worldwide. As crucial agronomic traits, leaves play an essential role in the growth of cotton plants, disease resistance, fiber quality, and yield. Pentatricopeptide repeat (PPR) proteins are a large family of nuclear-encoded proteins involved in organellar or nuclear RNA metabolism. Using a virus-induced gene silencing (VIGS) assay, we found that cotton plants displayed variegated yellow leaf phenotypes with decreased chlorophyll content when silencing the expression of the PPR gene named GhCTSF1. GhCTSF1 encodes a chloroplast-localized protein containing only two PPR motifs. Disruption of GhCTSF1 substantially reduced the splicing efficiency of rpoC1 intron 1 and ycf3 intron 2. Loss of function of the GhCTSF1 ortholog EMB1417 causes splicing defects in rpoC1 and ycf3-2, leading to impaired chloroplast structure and decreased photosynthetic rates in Arabidopsis. Moreover, we found that GhCTSF1 interacted with two splicing factors, GhCRS2 and GhWTF1. Defects in GhCRS2 and GhWTF1 severely affect the intron splicing of rpoC1 and ycf3-2 in cotton, leading to defects in chloroplast development and a reduction in photosynthesis. Our results suggest that GhCTSF1 is specifically required for splicing rpoC1 and ycf3-2 in cooperation with GhCRS2 and GhWTF1.