黑尾叶蝉PGRP6负调控共生菌抑制水稻矮缩病毒的经卵传播

doi:10.3864/j.issn.0578-1752.2025.05.007

Abstract

Abstract:

【Objective】Rice dwarf virus (RDV) causes rice dwarf disease, which is a significant threat to rice production in southern China. This study builds on prior research and employs molecular biology techniques to investigate the role of PGRP6 in regulating the vertical transmission of RDV from symbiotic bacteria within the vector insect Nephotettix cincticeps, thereby establishing a foundation for biological control strategies against rice viral diseases.【Method】Yeast two-hybrid technology was used to screen peptidoglycan recognition proteins (PGRPs) that interact with three proteins (prp, Nasuia porin, and P8) related to transovarial transmission, and the interaction was further confirmed by GST pull-down assay. Real-time fluorescence quantitative PCR technology was employed to assess the differential expression of related genes in nonviruliferous and viruliferous insects, as well as the changes in transcription levels of symbiotic bacteria following interference with PGRP6 and prp. Western blot experiments further confirmed the impact of RNAi treatment on symbiotic bacterial membrane proteins in insects. Following the reduction of PGRP6 expression by RNAi, variations were observed in the survival rate of N. cincticeps, the virus infection rate of the offspring, and the viral nucleic acid level of the infected offspring. Fluorescence in situ hybridization (FISH) and fluorescent immunolabeling techniques were employed to visualize the distribution of prp, PGRP6, Nasuia, and RDV. The functionality of the PGRP6 protein was assessed through inhibition zone assays and experiments on peptidoglycan degradation.【Result】PGRP6 interacted with prp, Nasuia porin, and P8. The expression levels of PGRP6 and prp were significantly elevated in RDV-infected N. cincticeps. Suppression of PGRP6 expression could facilitate the proliferation of symbiotic bacteria Nasuia and Sulcia, while inhibition of prp reduced the proliferation of symbiotic bacteria, acting oppositely to PGRP6. When both were interfered simultaneously, there was no significant effect on the proliferation of symbiotic bacteria. Treatment with dsPGRP6 significantly decreased the survival rate of N. cincticeps and enhanced the transmission of RDV to their offspring, resulting in a substantially higher viral load in the progeny compared to the control group. Immunofluorescence analysis revealed that PGRP6 co-localized with prp, Nasuia, and RDV within ovarian tissue, where prp and Nasuia exhibited a wrapping relationship. Ultimately, PGPR6 was identified as possessing antibacterial properties that inhibited the proliferation of symbiotic bacteria.【Conclusion】PGRP6 maintains the homeostasis of ovarian tissue in N. cincticeps by inhibiting the proliferation of symbiotic bacteria, which functionally antagonizes prp. This mechanism not only ensures the vitality of N. cincticeps but also regulates the transovarial transmission of RDV associated with bacterial symbionts.

Key words: rice dwarf virus (RDV), Nephotettix cincticeps, peptidoglycan recognition protein (PGRP), proline-rich protein (prp), symbiotic bacterium, ovary, transovarial transmission, antibacterial

XU YuanYuan, JIA DongSheng, BIN Yu, WEI TaiYun. PGRP6 Negatively Regulates Symbiotic Bacteria to Prevent the Transovarial Transmission of RDV in Nephotettix cincticeps[J].Scientia Agricultura Sinica, 2025, 58(5): 907-917.

Figures/Tables 5

Table 1

Primer sequences used in this study"

引物名称Primer name	序列Sequence (5′-3′)
AD-PGRP3 F	ccagattacgctcatATGGGTTTAGGTATTGATGAAAATATAAATACCTATTACAAAGT
AD-PGRP3 R	agctcgagctcgatgTTACGCAGCTTGTCTAAGATTCGGGT
AD-PGRP4 F	ccagattacgctcatATGGGGTTGATGGATAAACCACC
AD-PGRP4 R	agctcgagctcgatgCTAGTAATCACCATATTTCCTTCGGTATTCTTCT
AD-PGRP6 F	ccagattacgctcatATGGGATTGTGGTCTTCAAAAGAGG
AD-PGRP6 R	agctcgagctcgatgCTACTCATGCCCTAGAATGTCGC
AD-PGRP7 F	ccagattacgctcatATGTCCGCACCAGAACCAG
AD-PGRP7 R	agctcgagctcgatgTTAAAATAATTTTGTCTCCCCACGGAAAGACTTAACTG
AD-PGRP11 F	ccagattacgctcatATGGCAAGCCCCGTAGTG
AD-PGRP11 R	agctcgagctcgatgTTAAAACAAGTCGACTCTTTCAAAGTTCTGT
AD-PGRP-12 F	ccagattacgctcatATGGGGGACATACCAGATGCT
AD-PGRP-12 R	agctcgagctcgatgTTATTGTTTAGGCGTGATCTGTTGATTTGTTCG
AD-PGRP14 F	ccagattacgctcatATGGGGAAAGATAAGTACAACCCATTTAGG
AD-PGRP14 R	agctcgagctcgatgTTATGAATAGTAAGGGTGGCGAGTGG
AD-PGRP18 F	ccagattacgctcatATGAAACCTATGCATAAACCAATACAAATAGCAATGTTTCT
AD-PGRP18 R	agctcgagctcgatgTCAGTCCCACGGAGGCTCGG
BD-prp F	agaggaggacctgcatatggATGCGTTATCATAAGGACTT
BD-prp R	ttatgcggccgctgcTAGTTATGCGGCCGCTGCTTAAAAGTCTTCAAGCATTG
BD-Nasuia_porin F	gaggacctgcatatgATGTCAAATTCAATTTATTTTTTTTTTATAG
BD-Nasuia_porin R	ttatgcggccgctgcTTAATCTAATAATTTTAAAAATTTTTTATAATAAG
BD-RDV P8 F	gaggacctgcatatgATGTCACGCCAGATGTGGT
BD-RDV P8 R	ttatgcggccgctgcCTAATTTGGTCTATAGTATCTTCCAAATACGGCG
Q-EF1 F	CAGTGAGAGCCGTTTTGAG
Q-EF1 R	AGGGCATCTTGTCAGAGGGC
Q-prp F	ATGTAACCACTCGTGCACCC
Q-prp R	CAACATTTCCGTGTCGCACT
Q-PGRP6 F	AGCGACATTCTAGGGCATGA
Q-PGRP6 R	ACAGCAGTCAACGTAAAGGC
Q-Nasuia 16S F	GGGGAAAACCTCGCGTTATA
Q-Nasuia 16S R	CCACTGCTGCCTCTCGTAAG
Q-Sulcia 16S F	GGGGACTCTAATAAGACTGC
Q-Sulcia 16S R	CTGAGATCGGCTTTCTGGAT
PGEX-RDV-P8 F	aaatcggatctggttATGTCACGCCAGATGTGGTTAG
PGEX-RDV-P8 R	gtcagtcacgatgcgCTAATTTGGTCTATAGTATCTTCCA
PGEX-Nasuia_porin F	aaatcggatctggCGTGGATCCCCGATGTCAAATTCAATTTATTT TTTTTTTATAG
PGEX-Nasuia_porin R	gtcagtcacgatgcgGATGAATTCCGGTTAATCTAATAATTTTAAAAATTT TTTATAATAAG
PGEX-PGRP6 F	aaatcggatctggttATGGGATTGTGGTCTTCAAAAGAGG
PGEX-PGRP6 R	gtcagtcacgatgcgCTCATGCCCTAGAATGTCGC
PET-prp F	gatataccatgggcgATGGTGAAAGTTGGAACCTCTTACG
PET-prp R	gagcggccgcaagctGAAAAGCTTTGGTTTGGGAGCG
PET-PGRP6 F	ggtggacagcaaatgATGGGATTGTGGTCTTCAAAAGAGG
PET-PGRP6 R	gtgctcgagtgcggcCTACTCATGCCCTAGAATGTCGCT
T7-GFP-F	attctctagaagcttaatacgactcactatagggATGAGTAAAGGAGAAGAACTT
T7-GFP-R	attctctagaagcttaatacgactcactatagggTTATTTGTATAGTTCATCCATG
T7-PGRP6 F	attctctagaagcttaatacgactcactatagggTCTTTTCTGAGCGACGGTGT
T7-PGRP6 R	attctctagaagcttaatacgactcactatagggACTGTTCCTGGCTGTATTGC
T7-prp F	attctctagaagcttaatacgactcactatagggACCTCTTACGTGCCGATCAA
T7-prp R	attctctagaagcttaatacgactcactatagggTTTGGTTTGGGAGCGCTTTT

Table 1

Fig. 1

Fig. 2

Fig. 3

Fig. 4

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PGRP6 Negatively Regulates Symbiotic Bacteria to Prevent the Transovarial Transmission of RDV in Nephotettix cincticeps

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