Published online 18 October 2018 Nucleic Acids Research, 2018, Vol. 46, No. 21 11381–11395 doi: 10.1093/nar/gky897 Homodimerization regulates an endothelial specific signature of the SOX18 transcription factor Mehdi Moustaqil1, Frank Fontaine2, Jeroen Overman2, Alex McCann2, Timothy L. Bailey3, Paulina Rudolffi Soto1, Akshay Bhumkar1, Nichole Giles1, Dominic J.B. Hunter1,2, Yann Gambin1, Mathias Francois2,* and Emma Sierecki1,* Downloaded from https://academic.oup.com/nar/article-abstract/46/21/11381/5134331 by UNSW user on 05 December 2018 1EMBL Australia node in Single Molecule Science and School of Medical Sciences, The University of New South Wales, Sydney, NSW 2031, Australia, 2Institute for Molecular Bioscience, The University of Queensland, Brisbane, QLD 4072, Australia and 3Department of Pharmacology, School of Medicine, University of Nevada, Reno, NV 89557, USA
Received February 08, 2018; Revised September 17, 2018; Editorial Decision September 20, 2018; Accepted September 26, 2018
ABSTRACT namic control of gene transcription is particularly im- portant during development as cell lineages are estab- During embryogenesis, vascular development re- lished. In mammals, many members of the SOX SRY- lies on a handful of transcription factors that in- related High-Mobility Group (HMG) box family act as struct cell fate in a distinct sub-population of the central regulators of gene expression to govern cell fate endothelium (1). The SOXF proteins that comprise in a variety of key processes (4–7), such as vascular net- SOX7, 17 and 18, are molecular switches modulating work assembly (8), cartilage formation and sex determi- arterio-venous and lymphatic endothelial differenti- nation (9,10), neurogenesis (11), as well early stage devel- ation (2,3). Here, we show that, in the SOX-F fam- opment and embryonic stem cell pluripotency (12). These ily, only SOX18 has the ability to switch between crucial roles are highlighted by the fact that many muta- a monomeric and a dimeric form. We characterized tions in SOX genes cause severe congenital diseases in hu- the SOX18 dimer in binding assays in vitro, and us- mans, such as XY sex reversal (SRY), campomelic dyspla- sia (SOX9), Waardenburg–Hirschsprung syndrome (SOX8) ing a split-GFP reporter assay in a zebrafish model and anophthalmia–esophageal–genital syndrome (SOX2). system in vivo. We show that SOX18 dimerization is A prominent feature of the SOX proteins is the presence of driven by a novel motif located in the vicinity of the C- a 79 amino acids region which delineates the HMG-box, terminus of the DNA binding region. Insertion of this the DNA binding and bending domain. The HMG-box is motif in a SOX7 monomer forced its assembly into present in all groups of SOX proteins (A-H, 20 SOX) and a dimer. Genome-wide analysis of SOX18 binding lo- is classically used as a reference to align and compare these cations on the chromatin revealed enrichment for a proteins since this region is highly conserved (7). It is made SOX dimer binding motif, correlating with genes with up of 3 ␣-helixes, whereby ␣1and␣2 are involved with a strong endothelial signature. Using a SOX18 small DNA binding while ␣3 is involved in protein-protein inter- molecule inhibitor that disrupts dimerization, we re- actions (13). The HMG-box recognizes a heptameric con- / / / vealed that dimerization is important for transcrip- sensus sequence (5-A TA TCAAA TG-3) on the DNA. The activity of SOX proteins at these binding locations is tion. Overall, we show that dimerization is a specific modulated by varying the combinations of protein-protein feature of SOX18 that enables the recruitment of key interactions which can cause activation or repression of endothelial transcription factors, and refines the se- transcription (14–16). In addition to the HMG-box com- lectivity of the binding to discrete genomic locations mon to all SOX genes, individual subgroups possess other assigned to endothelial specific genes. functional domains that include: transactivation domain (TAD), coiled–coil, and proline-rich domains. The presence of these domains within the same group is likely to account INTRODUCTION for redundancy mechanism, an essential safety net to insure Understanding how transcription factors (TFs) orches- proper embryonic development (4). In particular, SOX pro- trate gene expression to instruct a phenotypic output teins within the F group (SOX7, SOX17 and SOX18) reg- is fundamental to biology and future therapeutics. Dy-
*To whom correspondence should be addressed: Emma Sierecki. Tel: +61 2 9385 8933; Fax: +61 2 9385 1389; Email: [email protected] Correspondence may also be addressed to Mathias Francois. Tel: +61 7 334 62494; Email: [email protected]