Biophysical Society Conference | Tahoe 2023

Proton Reactions: From Basic Science to Biomedical Applications

Tuesday Speaker Abstracts

HOW INTRACELLULAR PH DYNAMICS REGULATES CELL BEHAVIORS Diane Barber 1 ; 1 University of California, San Francisco, Department of Cell and Tissue Biology, San Francisco, CA, USA Intracellular pH (pHi) was previously thought to be constant for homeostasis but is now established to be dynamic, changing during cell cycle progression, cell migration, and cell differentiation to regulate myriad cell behaviors. Moreover, pHi is dysregulated in diseases, including being constitutively increased in cancers and decreased in neurodegenerative disorders. The molecular mechanisms mediating pHi-regulated cell behaviors, however, remain poorly understood. Our work bridges protein electrostatics and cell biology to reveal how pHi dynamics regulates cell behaviors through protonation of titrating amino acids as a post-translational modification modulating protein structure and function (Ann Rev Biophys. 42:289). I will review the design principles and functions of endogenous “pH sensors” controlling cell polarity and adhesion, actin assemblies, stem cell differentiation, and tumorigenesis. I also will present new findings on how pHi dynamics can regulate gene expression by modulating transcription factor DNA binding. We identified at least 65 transcription factors in diverse families having a conserved histidine, which in available structures forms a hydrogen bond with DNA nucleotides. FOX family transcription factors have an invariant DNA-binding histidine, and we confirmed using recombinant FOXM1 and FOXC2 and fluorescence anisotropy that pH regulates affinities for binding thymine compared with adenine, which are generally hydrogen bond acceptors and donors, respectively. We confirmed significance of the conserved histidine by showing pH independent differences in binding affinities for FOXC2-His122K and FOXC2-H122N, the latter being a recurrent mutation in cancers. We also confirmed pHi regulates FOXC2 activity in cells by using luciferase reporters with distinct DNA binding motifs. Our data identify pH dynamics as a previously unrecognized regulator of DNA-binding selectivity for FOXM1 and FOXC2. This regulatory mode has broad significance because histidine-DNA nucleotide binding is also reported for all members of the MITF/Myc family, most members of the SOX family, and selective members of STAT and ETS families.

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