The harpin protein HrpN_Ea induces Arabidopsis resistance to the green peach aphid by activating the ethylene signalling pathway and by recruiting EIN2, an essential regulator of ethylene signalling, for a defence response in the plant. We investigated 37 ethylene-inducible Arabidopsis transcription factor genes for their effects on the activation of ethylene signalling and insect defence. Twenty-eight of the 37 genes responded to both ethylene and HrpN_Ea, and showed either increased or inhibited transcription, while 18 genes showed increased transcription not only by ethylene but also by HrpN_Ea. In response to HrpN_Ea, transcription levels of 22 genes increased, with AtMYB44 being the most inducible, six genes had decreased transcript levels, and nine remained unchanged. When Arabidopsis mutants previously generated by mutagenicity at the 37 genes were surveyed, 24 mutants were similar to the wild type plant while four mutants were more resistant and nine mutants were more susceptible than wild type to aphid infestation. Aphid-susceptible mutants showed a greater susceptibility for atmyb15, atmyb38 and atmyb44, which were generated previously by T-DNA insertion into the exon region of AtMYB15 and the promoter regions of AtMYB38 and AtMYB44. The atmyb44 mutant was the most susceptible to aphid infestation and most compromised in induced resistance. Resistance accompanied the expression of PDF1.2, an ethylene signalling marker gene that requires EIN2 for transcription in wild type but not in atmyb15, atmyb38, and atmyb44, suggesting a disruption of ethylene signalling in the mutants. However, only atmyb44 incurred an abrogation in induced EIN2 expression, suggesting a close relationship between AtMYB44 and EIN2.

In Arabidopsis thaliana (Arabidopsis) treated with the harpin protein HrpN_Ea, resistance to the green peach aphid Myzus persicae, a generalist phloem-feeding insect, develops with induced expression of the AtMYB44 gene. Special GLUCAN SYNTHESIS-LIKE (GSL) genes and 𝛽-1,3-glucan callose play an important role in plant defence responses to attacks by phloem-feeding insects. Here we report that AtGLS5 and AtMYB44 are both required for HrpN_Ea-induced repression of M. persicae feeding from the phloem of Arabidopsis leaves. In 24 h successive surveys on large-scale aphid populations, the proportion of feeding aphids was much smaller in HrpN_Ea-treated plants than in control plants, and aphids preferred to feed from the 37 tested atgsl mutants rather than the wild-type plant. The atgsl mutants were generated previously by mutagenesis in 12 identified AtGSL genes (AtGSL1 through AtGSL12); in the 24 h survey, both atgsl5 and atgsl6 tolerated aphid feeding, and atgsl5 was the most tolerant. Consistently, atgsl5 was also most inhibitive to the deterrent effect of HrpN_Ea on the phloem-feeding activity of aphids as monitored by the electrical penetration graph technique. These results suggested an important role of the AtGSL5 gene in the effect of HrpN_Ea. In response to HrpN_Ea, AtGSL5 expression and callose deposition were induced in the wild-type plant but not in atgsl5. In response to HrpN_Ea, moreover, the AtMYB44 gene known to be required for repression of aphid reproduction on the plant was also required for repression of the phloem-feeding activity. Small amounts of the AtGSL5 transcript and callose deposition were detected in the atmyb44 mutant, as in atgsl5. Both mutants performed similarly in tolerating the phloem-feeding activity and impairing the deterrent effect of HrpN_Ea, suggesting that AtGSL5 and AtMYB44 both contributed to the effect.