High levels of salinity induce serious oxidative damage in plants. Flavonoids, as antioxidants, have important roles in reactive oxygen species (ROS) scavenging. In the present study, the tobacco R2R3 MYB type repressor, NtMYB4, was isolated and characterized. The expression of
NtMYB4
was suppressed by salinity. Overexpression of
NtMYB4
reduced the salt tolerance in transgenic tobacco plants. NtMYB4 repressed the promoter activity of
NtCHS1
and negatively regulated its expression. Rutin accumulation was significantly decreased in
NtMYB4
overexpressing transgenic plants and
NtCHS1
RNAi silenced transgenic plants. Moreover, high H
2
O
2
and
contents were detected in both types of rutin-reduced transgenic plants under high salt stress. In addition, exogenous rutin supplementation effectively scavenged ROS (H
2
O
2
and
) and improved the salt tolerance of the rutin-reduced transgenic plants. In contrast,
NtCHS1
overexpressing plants had increased rutin accumulation, lower H
2
O
2
and
contents, and higher tolerance to salinity. These results suggested that tobacco NtMYB4 acts as a salinity response repressor and negatively regulates
NtCHS1
expression, which results in the reduced flavonoid accumulation and weakened ROS-scavenging ability under salt stress.
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