Red fruit peel is one of pear's most valuable economic traits and is mainly determined by anthocyanins.Many pear cultivars with a red peel originated from bud sports;however,little is known about the genetic mecha...Red fruit peel is one of pear's most valuable economic traits and is mainly determined by anthocyanins.Many pear cultivars with a red peel originated from bud sports;however,little is known about the genetic mechanisms underlying this trait.We have previously identified a mutant Pp BBX24 containing a 14-nucleotide deletion in the coding region(Ppbbx24-del)as the only known variant associated with the red coloration of the mutant‘Red Zaosu'pear(Pyrus pyrifolia White Pear Group).Herein,we analyzed the role of the mutant gene in red coloration and its mechanism of action.The results showed that light promoted red peel coloration in the‘Red Zaosu'pear,and Ppbbx24-del positively affected light-induced anthocyanin biosynthesis,while normal Pp BBX24 had the opposite effects.Transient and stable transformation experiments confirmed that Ppbbx24-del could promote anthocyanin accumulation in pear fruit peels,calli,and tobacco flowers.Due to the loss of nuclear localization sequence(NLS)and viral protein(VP)domains,Ppbbx24-del co-localized in the nucleus and cytoplasm,whereas PpBBX24 localized only in the nucleus.Real-time PCR and transcriptome analyses indicated that PpM YB10 and PpH Y5 are highly expressed in the‘Red Zaosu'pear.In yeast one-hybrid and dual-luciferase assays,Ppbbx24-del and PpHY5 independently promoted the expression of PpC HS,PpC HI,and PpM YB10 by binding to their promoters;however,PpBBX24 did not affect the expression of these genes.Additionally,we found that Ppbbx24-del and PpHY5 had additive effects on the expression of PpC HS,PpC HI,and PpMYB10,as they promote the expression of anthocyanin synthesis genes separately.The co-expression of PpB BX24 and PpHY5 inhibited the activation of downstream genes by PpHY5,which was attributed to the interaction between the two loci.In conclusion,our results clarify the molecular mechanism by which mutant Ppbbx24-del and PpBBX24 exert opposite effects in regulating anthocyanin accumulation in pear.These findings lay an important theoretical foundation for using Ppbbx24-del to create red pear cultivars.展开更多
Red skin is an important quality trait for pear fruits and is determined by the concentration and composition of anthocyanins.The regulatory mechanism underlying anthocyanin accumulation is a popular topic in fruit re...Red skin is an important quality trait for pear fruits and is determined by the concentration and composition of anthocyanins.The regulatory mechanism underlying anthocyanin accumulation is a popular topic in fruit research.Red mutants are ideal materials for studying the molecular mechanism of color diversity in pear.Although several red pear mutants have been cultivated and are in production,no exact locus containing the responsible genetic mutation has been identified.In this study,by combining the bulked segregant analysis with whole-genome sequencing,we identified a 14 nucleotide deletion mutation in the coding region of the PpBBX24 gene from the red pear mutant“Zaosu Red”.We further verified that the deletion was present only in the red mutant of“Zaosu”and in its red offspring,which was different from that which occurred in other red pear fruits.This deletion results in a coding frame shift such that there is an early termination of the PpBBX24 gene and loss of key NLS and VP domains from PpBBX24.The lost domains may reduce or alter the normal function of PpBBX24.In addition,we found that the transcript levels of the PpMYB10 and PpHY5 genes in red samples were significantly higher than those in green samples,whereas the results for the normal-type PpBBX24 gene were the opposite.We ultimately revealed that the 14 nucleotide deletion mutation in the coding region of the PpBBX24 gene is associated with the red skin of the“Zaosu Red”pear.This finding of somatic mutational events will be helpful for breeding new red pear cultivars and for understanding the regulatory mechanisms involved in pear skin pigmentation.展开更多
基金National Natural Science Foundation of China(32072531)the Agricultural Science and Technology Innovation Program of the Chinese Academy of Agricultural Sciences(CAAS-ASTIP-2016-RIP)+2 种基金the National Key Research and Development Program of China(2021YFE0190700)the ScienceTechnology&Innovation Funding Authority(STDF)of Egypt(43093)for funding this work。
文摘Red fruit peel is one of pear's most valuable economic traits and is mainly determined by anthocyanins.Many pear cultivars with a red peel originated from bud sports;however,little is known about the genetic mechanisms underlying this trait.We have previously identified a mutant Pp BBX24 containing a 14-nucleotide deletion in the coding region(Ppbbx24-del)as the only known variant associated with the red coloration of the mutant‘Red Zaosu'pear(Pyrus pyrifolia White Pear Group).Herein,we analyzed the role of the mutant gene in red coloration and its mechanism of action.The results showed that light promoted red peel coloration in the‘Red Zaosu'pear,and Ppbbx24-del positively affected light-induced anthocyanin biosynthesis,while normal Pp BBX24 had the opposite effects.Transient and stable transformation experiments confirmed that Ppbbx24-del could promote anthocyanin accumulation in pear fruit peels,calli,and tobacco flowers.Due to the loss of nuclear localization sequence(NLS)and viral protein(VP)domains,Ppbbx24-del co-localized in the nucleus and cytoplasm,whereas PpBBX24 localized only in the nucleus.Real-time PCR and transcriptome analyses indicated that PpM YB10 and PpH Y5 are highly expressed in the‘Red Zaosu'pear.In yeast one-hybrid and dual-luciferase assays,Ppbbx24-del and PpHY5 independently promoted the expression of PpC HS,PpC HI,and PpM YB10 by binding to their promoters;however,PpBBX24 did not affect the expression of these genes.Additionally,we found that Ppbbx24-del and PpHY5 had additive effects on the expression of PpC HS,PpC HI,and PpMYB10,as they promote the expression of anthocyanin synthesis genes separately.The co-expression of PpB BX24 and PpHY5 inhibited the activation of downstream genes by PpHY5,which was attributed to the interaction between the two loci.In conclusion,our results clarify the molecular mechanism by which mutant Ppbbx24-del and PpBBX24 exert opposite effects in regulating anthocyanin accumulation in pear.These findings lay an important theoretical foundation for using Ppbbx24-del to create red pear cultivars.
基金the National Key Research and Development Program of China(2018YFD1000102)the Agricultural Science and Technology Innovation Program of the Chinese Academy of Agricultural Sciences(CAAS-ASTIP-2016-RIP)programs for funding this work.
文摘Red skin is an important quality trait for pear fruits and is determined by the concentration and composition of anthocyanins.The regulatory mechanism underlying anthocyanin accumulation is a popular topic in fruit research.Red mutants are ideal materials for studying the molecular mechanism of color diversity in pear.Although several red pear mutants have been cultivated and are in production,no exact locus containing the responsible genetic mutation has been identified.In this study,by combining the bulked segregant analysis with whole-genome sequencing,we identified a 14 nucleotide deletion mutation in the coding region of the PpBBX24 gene from the red pear mutant“Zaosu Red”.We further verified that the deletion was present only in the red mutant of“Zaosu”and in its red offspring,which was different from that which occurred in other red pear fruits.This deletion results in a coding frame shift such that there is an early termination of the PpBBX24 gene and loss of key NLS and VP domains from PpBBX24.The lost domains may reduce or alter the normal function of PpBBX24.In addition,we found that the transcript levels of the PpMYB10 and PpHY5 genes in red samples were significantly higher than those in green samples,whereas the results for the normal-type PpBBX24 gene were the opposite.We ultimately revealed that the 14 nucleotide deletion mutation in the coding region of the PpBBX24 gene is associated with the red skin of the“Zaosu Red”pear.This finding of somatic mutational events will be helpful for breeding new red pear cultivars and for understanding the regulatory mechanisms involved in pear skin pigmentation.
