TY - JOUR

T1 - Architecture of DNA elements mediating ARF transcription factor binding and auxin-responsive gene expression in Arabidopsis

AU - Freire-Rios, Alejandra

AU - Tanaka, Keita

AU - Crespo, Isidro

AU - Van der Wijk, Elmar

AU - Sizentsova, Yana

AU - Levitsky, Victor

AU - Lindhoud, Simon

AU - Fontana, Mattia

AU - Hohlbein, Johannes

AU - Roeland Boer, D.

AU - Mironova, Victoria

AU - Weijers, Dolf

N1 - Copyright © 2020 the Author(s). Published by PNAS.

PY - 2020/9/29

Y1 - 2020/9/29

N2 - The hormone auxin controls many aspects of the plant life cycle by regulating the expression of thousands of genes. The transcriptional output of the nuclear auxin signaling pathway is determined by the activity of AUXIN RESPONSE transcription FACTORs (ARFs), through their binding to cis-regulatory elements in auxinresponsive genes. Crystal structures, in vitro, and heterologous studies have fueled a model in which ARF dimers bind with high affinity to distinctly spaced repeats of canonical AuxRE motifs. However, the relevance of this "caliper" model, and the mechanisms underlying the binding affinities in vivo, have remained elusive. Here we biochemically and functionally interrogate modes of ARF-DNA interaction. We show that a single additional hydrogen bond in Arabidopsis ARF1 confers high-affinity binding to individual DNA sites. We demonstrate the importance of AuxRE cooperativity within repeats in the Arabidopsis TMO5 and IAA11 promoters in vivo. Meta-analysis of transcriptomes further reveals strong genome-wide association of auxin response with both inverted (IR) and direct (DR) AuxRE repeats, which we experimentally validated. The association of these elements with auxininduced up-regulation (DR and IR) or down-regulation (IR) was correlated with differential binding affinities of A-class and B-class ARFs, respectively, suggesting a mechanistic basis for the distinct activity of these repeats. Our results support the relevance of highaffinity binding of ARF transcription factors to uniquely spaced DNA elements in vivo, and suggest that differential binding affinities of ARF subfamilies underlie diversity in cis-element function.

AB - The hormone auxin controls many aspects of the plant life cycle by regulating the expression of thousands of genes. The transcriptional output of the nuclear auxin signaling pathway is determined by the activity of AUXIN RESPONSE transcription FACTORs (ARFs), through their binding to cis-regulatory elements in auxinresponsive genes. Crystal structures, in vitro, and heterologous studies have fueled a model in which ARF dimers bind with high affinity to distinctly spaced repeats of canonical AuxRE motifs. However, the relevance of this "caliper" model, and the mechanisms underlying the binding affinities in vivo, have remained elusive. Here we biochemically and functionally interrogate modes of ARF-DNA interaction. We show that a single additional hydrogen bond in Arabidopsis ARF1 confers high-affinity binding to individual DNA sites. We demonstrate the importance of AuxRE cooperativity within repeats in the Arabidopsis TMO5 and IAA11 promoters in vivo. Meta-analysis of transcriptomes further reveals strong genome-wide association of auxin response with both inverted (IR) and direct (DR) AuxRE repeats, which we experimentally validated. The association of these elements with auxininduced up-regulation (DR and IR) or down-regulation (IR) was correlated with differential binding affinities of A-class and B-class ARFs, respectively, suggesting a mechanistic basis for the distinct activity of these repeats. Our results support the relevance of highaffinity binding of ARF transcription factors to uniquely spaced DNA elements in vivo, and suggest that differential binding affinities of ARF subfamilies underlie diversity in cis-element function.

KW - ARF transcription factors

KW - Auxin

KW - Plant biology

KW - Protein-DNA interaction

KW - Transcriptional regulation

KW - auxin

KW - SPECIFICITY

KW - MECHANISM

KW - INITIATION

KW - protein-DNA interaction

KW - BOX PROTEIN TIR1

KW - plant biology

KW - FAMILY

KW - PERCEPTION

KW - TRANSPORT

KW - EVOLUTION

KW - regulation

KW - REPRESSION

KW - transcriptional

KW - PROMOTER

UR - http://www.scopus.com/inward/record.url?scp=85092428276&partnerID=8YFLogxK

U2 - 10.1073/pnas.2009554117

DO - 10.1073/pnas.2009554117

M3 - Article

C2 - 32929017

AN - SCOPUS:85092428276

VL - 117

SP - 24557

EP - 24566

JO - Proceedings of the National Academy of Sciences of the United States of America

JF - Proceedings of the National Academy of Sciences of the United States of America

SN - 0027-8424

IS - 39

ER -