TWIST1 Homodimers and Heterodimers Orchestrate Lineage-Specific Differentiation

Abstract

The extensive array of basic helix-loop-helix (bHLH) transcription factors and their combinations as dimers underpin the diversity of molecular function required for cell type specification during embryogenesis. The bHLH factor TWIST1 plays pleiotropic roles during development. However, which combinations of TWIST1 dimers are involved and what impact each dimer imposes on the gene regulation network controlled by TWIST1 remain elusive. In this work, proteomic profiling of human TWIST1-expressing cell lines and transcriptome analysis of mouse cranial mesenchyme have revealed that TWIST1 homodimers and heterodimers with TCF3, TCF4, and TCF12 E-proteins are the predominant dimer combinations. Disease-causing mutations in TWIST1 can impact dimer formation or shift the balance of different types of TWIST1 dimers in the cell, which may underpin the defective differentiation of the craniofacial mesenchyme. Functional analyses of the loss and gain of TWIST1–E-protein dimer activity have revealed previously unappreciated roles in guiding lineage differentiation of embryonic stem cells: TWIST1–E-protein heterodimers activate the differentiation of mesoderm and neural crest cells, which is accompanied by the epithelial-to-mesenchymal transition. At the same time, TWIST1 homodimers maintain the stem cells in a progenitor state and block entry to the endoderm lineage.

TWIST1 and chromatin regulatory proteins interact to guide neural crest cell differentiation

“…Differential binding analysis of TWIST1 using sum-normalized peptide-spectrum match (PSM) values ( Figure 1—figure supplement 2B,C ; see Materials and methods) revealed 140 putative TWIST1 interactors in NCCs (p<0.05; Fold-change >3; PSM#>2; Figure 1B , Supplementary file 1 ). These candidates included 4 of 56 known TWIST1 interactors, including TCF3, TCF4, TCF12, and GLI3 (overlap odds ratio = 18.05, Chi-squared test p-value=0.0005; Agile Protein Interactomes DataServer [APID]) ( Alonso-López et al, 2019 ; Fan et al, 2020 ). Despite that the APID database covers a broad spectrum of protein interaction in different cell line models, it was noted that the TWIST1 partners, TCF3, 4, and 12 that were recurrently identified in yeast-two-hybrid, immunoprecipitation and in vitro interaction assays were recovered by BioID ( El Ghouzzi et al, 2000 ; Firulli et al, 2007 ; Fu et al, 2011 ; Teachenor et al, 2012 ; Sharma et al, 2013 ; Kotlyar et al, 2015 ; Li et al, 2015 ).…”

PGC1α Loss Promotes Lung Cancer Metastasis through Epithelial-Mesenchymal Transition

“…TWIST1, EMT-promoting transcription factor, is an essential for cancer metastasis [ 49 ]. A recent report revealed that the TWIST1 homodimer and heterodimer interactions with E-proteins, such as TCF3/E2A, TCF4/ITF-2, and TCF12/HEB, enhances EMT [ 50 ]. Among those, the class I bHLH factor, TCF4, is one of the last additions among the EMT regulating factors, and acts as an E-cadherin repressor by interacting with other molecules.…”

Mechanisms of Binding Specificity among bHLH Transcription Factors

“…Finally, certain post-translational modifications can also affect DNA binding of bHLH factors. Phosphorylation of Serines or Threonines in the basic (e.g., HAND, Hes1, NeuroM (NeuroD4) and myogenic factors) [101][102][103][104], HLH (e.g., Twist1, HAND, and Neurogenin/Achaete-Scute/Atonal subfamilies) [70,101,105,106] and other (e.g., Max) [107] domains can impede DNA binding of the mentioned factors.…”