Papaya CpWRI3 is Involved in the Regulation of Fatty Acid Accumulation During Fruit Postharvest

doi:10.16420/j.issn.0513-353x.2022-0590

Acta Horticulturae Sinica ›› 2023, Vol. 50 ›› Issue (8): 1637-1648.doi: 10.16420/j.issn.0513-353x.2022-0590

• Genetic & Breeding·Germplasm Resources·Molecular Biology • Previous Articles Next Articles

Papaya CpWRI3 is Involved in the Regulation of Fatty Acid Accumulation During Fruit Postharvest

MO Yuxing, ZHOU Yan, LIN Runting, DUO Yongxing, ZHANG Tao, LIU Kaidong^**()

Life Science and Technology School of Lingnan Normal University，Zhanjiang，Guangdong 524048，China

Received:2022-08-26 Revised:2023-01-18 Online:2023-08-25 Published:2023-08-23
Contact: LIU Kaidong

Abstract

Abstract:

A WRI family regulator gene named CpWRI3 was cloned from papaya fruit. Amino acid sequence alignment and evolutionary analyses showed that CpWRI3 has high homology with AtWRI3/4. Transcriptional factor activity analysis showed that CpWRI3 has transcriptional activate function. The expression level of CpWRI3 increased during the fruit ripening process，peaked at eight days after harvested，indicating that CpWRI3 plays an important role in the late stage of fruit ripening. In order to explore the role of CpWRI3 in regulating fatty acid biosynthesis，transient overexpression of CpWRI3 was performed. The data showed that overexpression of CpWRI3 significantly induced the contents of lauric acid，palmitoleic acid and linoleic acid in papaya fruit. In vitro Electrophoretic Mobility Shift Assay（EMSA）confirmed that CpWRI3 could specifically bind to AW-box elements on promoters of downstream target genes CpKAS1 and CpACC1. Using luciferase system，CpWRI3 can induce the promoter activities of CpKAS1 and CpACC1 genes in plants. These results revealed that CpWRI3 may induce the biosynthesis of lauric acid，palmitoleic acid and linoleic acid.

Key words: papaya, fruit, ripening, fatty acid, transcription regulation, WRI family

MO Yuxing, ZHOU Yan, LIN Runting, DUO Yongxing, ZHANG Tao, LIU Kaidong. Papaya CpWRI3 is Involved in the Regulation of Fatty Acid Accumulation During Fruit Postharvest[J]. Acta Horticulturae Sinica, 2023, 50(8): 1637-1648.

Figures/Tables 11

Table 1 Primer sequences

引物用途 Purpose	引物名称 Primier name	引物序列（5′-3′） Sequence
CpWRI3基因扩增 Amplification of CpWRI3 gene	CpWRI3-F	CAATGCACAGAATGGTGCTGG
CpWRI3基因扩增 Amplification of CpWRI3 gene	CpWRI3-R	TTGAAACATGGTCGGCACGAG
荧光定量检测引物 Real-time PCR of CpWRI3	CpWRI3-RT-F	GCACATACGCAACGCAAGAA
荧光定量检测引物 Real-time PCR of CpWRI3	CpWRI3-RT-R	TTCGGGATTAGGGTTTGGCG
荧光定量内参引物 Real-time PCR of CpActin	CpActin-RT-F	CTGTGCATGATGACTGCAAG
荧光定量内参引物 Real-time PCR of CpActin	CpActin-RT-R	TCATCTTGCTTCTCACCTTG

Table 2 Primer sequences of Real-time PCR

目的基因 Gene	引物序列（5′-3′） Sequence
CpWRI3	F：TGCTGGAACGAATCGCAGAA;R：TCGTGCAACGCCTCTGTATT
CpKAS1	F：TGCGCATGCAACTTCAACTC;R：AACGTTCACCTCATGTCGCT
CpACP2	F：AAGTATGGTGATCGGGTGGC;R：TCCTGATCCCCTCACCACAT
CpBCCP2	F：GCGTTGGCTCAACCACAATC;R：CGACCTTCACAAAGGGTGGT
CpACC1	F：TAGCTGTGCGTGTGACAAGT;R：TTGGTTCGGATTTCTCCCCG

Fig. 1 The PCR product of CpWRI3 gene Marker：DL5 000.

Fig. 2 3D structure of CpWRI3 protein

Fig. 3 Alignment of the predicted CpWRI3 protein between papaya and Arabidopsis thaliana（A）and the phylogenetic tree of CpWRI3 protein and WRI proteins from other species（B） The data of B on the branch represents the bootstrap value.

Fig. 4 Transcriptional activity of CpWRI3 Student’s t-test，*** P < 0.001.

Fig. 5 Expression analysis of CpWRI3 gene during the fruit ripening process

Fig. 6 The expression change of CpWRI3 and relaive genesin CpWRI3 transient overexpressing（OE）and the control fruit Student’s t-test. **P < 0.01;***P < 0.001.

Fig. 7 The changes of content of lauric acid，palmitoleic acid，and linoleic acidin CpWRI3 transient overexpressing（OE）and the control fruit Student’s t-test. **P < 0.01;***P < 0.001.

Fig. 8 CpWRI3 protein was specifically binding to the AW-box in promoter of CpKAS1 and CpACC1 genes

Fig. 9 Vectors used in transient expression analysis（A）and relative LUC/REN ratio（B）

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Papaya CpWRI3 is Involved in the Regulation of Fatty Acid Accumulation During Fruit Postharvest

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