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. 2019 Jul 16;31(9):2089–2106. doi: 10.1105/tpc.18.00991

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© 2019 American Society of Plant Biologists. All rights reserved.

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Figure 8. — FHY3 Interacts with MYC2 to Coordinately Regulate LOX2 Expression.

(A) Yeast two-hybrid assay shows that FHY3 interacts with MYC2, MYC3, and MYC4. FHY3 was fused with the LexA DNA binding domain in pEG202. MYC2, MYC3, and MYC4 were individually fused with the activation domain (AD) in pB42AD.

(B) Firefly LCI assay shows that FHY3 interacts with MYC2 in plant cells. N. benthamiana leaves were transformed with the construct pairs cLUC-FHY3/nLuc, cLUC-FHY3/nLuc-MYC2, and cLuc/nLuc-MYC2.

(C) Firefly LCI assay shows that FHY3 interacts with MYC3 in plant cells. N. benthamiana leaves were transformed with the construct pairs cLUC-FHY3/nLuc, cLUC-FHY3/nLuc-MYC3, and cLuc/nLuc-MYC3.

(D) Co-IP assay showing that FHY3 associates with MYC2 in planta. Protein extracts from 6-d-old seedlings expressing FLAG-FHY3-HA and MYC2-4myc were immunoprecipitated using anti-FLAG antibody and blotted with anti-FLAG or anti-MYC antibody, respectively.

(E) and (F) Transient expression assay shows that FHY3 and MYC2 coordinately regulate ProLOX2:LUC expression. The N. benthamiana leaves were infiltrated with A. tumefaciens transformed with the indicated reporter and effector constructs. (E) shows a representative image of luciferase activity in an N. benthamiana leaf. (F) shows quantification of the transient expression assay. **, P < 0.01, Student’s t test. Values are means ± sd, n = 3.

(G) and (H) Transient expression assay shows that JAZ1 represses the transcriptional activity of FHY3 on ProLOX2:LUC. (G) shows a representative image of luciferase activity in an N. benthamiana leaf. (H) shows quantification of the transient expression assay. **, P < 0.01, Student’s t test. Values are means ± sd, n = 3.

(I) qRT-PCR analysis of LOX2 expression in Col, fhy3-11 far1-4, myc2-2, and fhy3-11 far1-4 myc2-2 plants grown under simulated shade. *, P < 0.05 and **, P < 0.01, Student’s t test. Data are means ± sd, n = 3.

(J) qRT-PCR analysis of LOX2 expression in Col, fhy3-11, MYC2OE, and MYC2OE fhy3-11 seedlings grown under simulated shade. **, P < 0.01, Student’s t test. Data are means ± sd, n = 3.