Summary Multi‐functional micro RNA s (mi RNA s) are emerging as key modulators of plant–pathogen interactions. Although the involvement of some mi RNA s in plant–insect interactions has been revealed, the underlying mechanisms are still elusive. The brown planthopper ( BPH ) is the most notorious rice ( Oryza sativa )‐specific insect that causes severe yield losses each year and requires urgent biological control. To reveal the mi RNA s involved in rice– BPH interactions, we performed mi RNA sequencing and identified BPH ‐responsive OsmiR396. Sequestering OsmiR396 by overexpressing target mimicry ( MIM 396) in three genetic backgrounds indicated that OsmiR396 negatively regulated BPH resistance. Overexpression of one BPH ‐responsive target gene of OsmiR396, growth regulating factor 8 ( Os GRF 8 ), showed resistance to BPH . Furthermore, the flavonoid contents increased in both the OsmiR396‐sequestered and the Os GRF 8 overexpressing plants. By analysing 39 natural rice varieties, the elevated flavonoid contents were found to correlate with enhanced BPH resistance. Artificial applications of flavonoids to wild type ( WT ) plants also increased resistance to BPH . A BPH ‐responsive flavanone 3‐hydroxylase ( OsF3H ) gene in the flavonoid biosynthetic pathway was proved to be directly regulated by Os GRF 8. A genetic functional analysis of OsF3H revealed its positive role in mediating both the flavonoid contents and BPH resistance. And analysis of the genetic correlation between OsmiR396 and OsF3H showed that down‐regulation of OsF3H complemented the BPH resistance characteristic and simultaneously decreased the flavonoid contents of the MIM 396 plants. Thus, we revealed a new BPH resistance mechanism mediated by the OsmiR396–Os GRF 8–OsF3H–flavonoid pathway. Our study suggests potential applications of mi RNA s in BPH resistance breeding.

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