UC San Diego

Spalt-Like Transcription Factor 1 (Sall1) is a critical regulator of microglia identity. Despite its importance, the transcriptional function of SALL1 and mechanisms regulating its expression are not fully understood. Here, we demonstrate that Sall1 is physically connected to a microglia-specific, environment-dependent super enhancer (SE) containing conserved binding sites for SMAD TFs downstream of TGF-beta signaling. Deletion of the SE in mice (EKO) led to selective loss of Sall1 transcript and SALL1 protein in microglia, resulting in downregulation of microglial identity genes and activation of inflammation- and aging-associated genes. Using ChIP-seq, we defined binding sites of SALL1 and leveraged EKO mice to probe how SALL1 shapes the regulatory landscape of microglia. We found thousands of putative enhancers whose activity was increased or abrogated by loss of SALL1; we further classified these enhancers as being ‘directly’ or ‘indirectly’ regulated by SALL1 based on overlap with SALL1 binding sites. Unexpectedly, motifs for SMADs are enriched within enhancers predicted to be directly activated by SALL1, suggesting that collaborative interactions between SALL1 and SMADs are required to establish microglia-specific gene expression. To test this hypothesis, we generated a conditional KO of the common co-SMAD Smad4 and determined the binding sites of SMAD4 in WT and EKO microglia. These studies demonstrated that SMAD4 binds to the Sall1 SE and controls Sall1 expression, and that its ability to activate other major microglia genes is dependent on co-binding with SALL1. Collectively, these results suggest a molecular basis for many of the transcriptional characteristics of yolk sac-derived microglia.