Abstract
Basal tumor propagating cells (TPCs) control squamous cell carcinoma (SCC) growth by self-renewing and differentiating into supra-basal SCC cells, which lack proliferative potential. While transcription factors such as SOX2 and KLF4 can drive these behaviors, their molecular roles and regulatory interactions with each other have remained elusive. Here, we show that PITX1 is specifically expressed in TPCs, where it co-localizes with SOX2 and TRP63 and determines cell fate in mouse and human SCC. Combining gene targeting with chromatin immunoprecipitation sequencing (ChIP-seq) and transcriptomic analyses reveals that PITX1 cooperates with SOX2 and TRP63 to sustain an SCC-specific transcriptional feed-forward circuit that maintains TPC-renewal, while inhibiting KLF4 expression and preventing KLF4-dependent differentiation. Conversely, KLF4 represses PITX1, SOX2, and TRP63 expression to prevent TPC expansion. This bi-stable, multi-input network reveals a molecular framework that explains self-renewal, aberrant differentiation, and SCC growth in mice and humans, providing clues for developing differentiation-inducing therapeutic strategies.