Kogan, L. | Search bioRxiv

bioRxiv2026

HCF1 orchestrates O-GlcNAcylation and affinity-dependent transcription through extended molecular determinants and register-shifted binding

We recently identified Host Cell Factor 1 (HCF1), a transcriptional co-regulator discovered more than thirty years ago, as a cancer dependency. To further understand its molecular functions and expand its known interactome, here, we screened a proteome-wide library of candidate HCF1 binding peptides and identified previously uncharacterized canonical and non-canonical HCF1-binding partners. Through deep mutational scanning (DMS) screening, we uncovered an extended set of molecular determinants of binding and show how mutations outside its previously established interacting residues impact binding affinity. Next, we uncover non-canonical HCF1 binders with an extended register-shifted two-amino acid sequence between their "anchor" histidine and tyrosine amino-acid residues, which we show critically contributes, in an affinity-dependent manner, to the downstream transcriptional activity of IRF1. Our data also shows that HCF1 promotes O-GlcNAcylation of the majority of its transcriptional binders. Overall, our results significantly expand the number and diversity of HCF1 binders and propose an enhanced mechanistic understanding of how HCF1 orchestrates transcription and O-GlcNAcylation.

molecular biology↗

bioRxiv2026

Uncovering cancer dependencies in peptide-interacting protein pockets

Cancer cells often become dependent on specific molecular functions. As many proteins perform multiple functions mediated by different pockets and interfaces, we hypothesized that we could identify distinct cancer dependencies and therapeutic vulnerabilities by disrupting peptide-binding pockets. To test this hypothesis, we screened a proteome-wide library of 7152 genetically encoded peptides across nine cancer cell lines. We identify common and selective dependencies on peptide-binding pockets and find that gene knockout and peptide-mediated inhibition of pockets often drive divergent phenotypes. For the common-essential gene HCF1, we identify a therapeutic window by using inhibitory peptides with varying affinity. Moreover, peptides targeting TLE1-4 reveal a dependency hidden in genetic screens by homolog redundancy. We also uncover that peptides inhibiting cyclin D drive specific suppression of leukemia cell proliferation and demonstrate that these peptides improve the potency of CDK4/6 inhibitors. Overall, our screening platform facilitates data-driven prioritization of molecular pockets for subsequent therapeutic translation.

cancer biology↗