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Blood orange is generally recognized to accumulate anthocyanins in its fruit pulp in a cold‐inducible manner. We observed that the fruit peel of blood orange can also accumulate anthocyanins under ample light conditions. Interestingly, purple pummelo can accumulate anthocyanins only in its fruit peel but not in its pulp. The mechanism underlying the tissue specificity of anthocyanin accumulation in citrus is unknown. Here, we show that the active promoter of Ruby1, a key activator of anthocyanin biosynthesis, is also light inducible in addition to its already known cold inducibility in blood orange. Electrophoretic mobility shift assays and transient expression assays showed that HY5 positively regulated the transcription of Ruby1 by binding to the G‐box motif (CACGTC). The tissue specificity of anthocyanin accumulation in the peel of purple pummelo may be due to the lack of a low temperature responsive element and a MYC binding site, which were shown to be involved in cold inducibility of CsRuby1 in blood orange by insertion of a long terminal repeat type retrotransposon in the promoter. These results bring new insights into the regulatory mechanism of anthocyanin biosynthesis in response to environmental stimuli and provide cis‐elements for genetic improvement of anthocyanin‐stable fruits rich in antioxidant metabolites. The accumulation of anthocyanins in fruit peel of blood orange in response to light is regulated by the G‐box (CACGTC) within the promoter of CsRuby1. On the other hand, cold‐inducible accumulation of anthocyanins in blood orange is regulated by the LTR region of the retrotransposon‐containing promoter of CsRuby1. The accumulation of anthocyanins in fruit peel of blood orange in response to light is regulated by the G‐box (CACGTC) within the promoter of CsRuby1. On the other hand, cold inducible accumulation of anthocyanins in blood orange is regulated by the LTR region of the retrotransposon‐containing promoter of CsRuby1. [ABSTRACT FROM AUTHOR]

Summary: Long intergenic non‐coding RNAs (lincRNAs) play important roles in various biological processes in plants. However, little information is known about the evolutionary characteristics of lincRNAs among closely related plant species. Here, we present a large‐scale comparative study of lincRNA transcription patterns in nine citrus species. By strand‐specific RNA‐sequencing, we identified 18 075 lincRNAs (14 575 lincRNA loci) from 34 tissue samples. The results indicated that the evolution of lincRNA transcription is more rapid than that of mRNAs. In total, 82.8–97.6% of sweet orange (Citrus sinensis) lincRNA genes were shown to have homologous sequences in other citrus genomes. However, only 15.5–28.8% of these genes had transcribed homologous lincRNAs in these citrus species, presenting a strong contrast to the high conservation of mRNA transcription (81.6–84.7%). Moreover, primitive and modern citrus lincRNAs were preferentially expressed in reproductive and vegetative organs, respectively. Evolutionarily conserved lincRNAs showed higher expression levels and lower tissue specificity than species‐specific lincRNAs. Notably, we observed a similar tissue expression pattern of homologous lincRNAs in sweet orange and pummelo (Citrus grandis), suggesting that these lincRNAs may be functionally conserved and selectively maintained. We also identified and validated a lincRNA with the highest expression in fruit that acts as an endogenous target mimic (eTM) of csi‐miR166c, and two lincRNAs that act as a precursor and target of csi‐miR166c, respectively. These lincRNAs together with csi‐miR166c could form an eTM166‐miR166c‐targeted lincRNA regulatory network that possibly affects citrus fruit development. Significance Statement: Here, 18 075 lincRNAs have been identified from 34 tissue samples from nine citrus species. The data uncover the rapid turnover of lincRNA transcription among different citrus species and highlights the potential contribution of the eTM166‐miR166‐targeted lincRNA regulatory network to fruit development. [ABSTRACT FROM AUTHOR]