Wild-type and L317P mutant ARHGAP36 isoform 2 interactome

ABSTRACT: ARHGAP36 is an atypical Rho GTPase-activating protein (GAP) family member that drives both spinal cord development and tumorigenesis, acting in part through an N-terminal motif that suppresses protein kinase A and activates Gli transcription factors. ARHGAP36 also contains isoform-specific N-terminal sequences, a central GAP-like module, and a unique C-terminal domain, and the functions of these regions remain unknown. Here we have mapped the ARHGAP36 structure-activity landscape using a deep sequencing-based mutagenesis screen and truncation mutant analyses. Using this approach, we have discovered several residues in the GAP homology domain that are essential for Gli activation and a role for the C-terminal domain in counteracting an N-terminal autoinhibitory motif that is present in certain ARHGAP36 isoforms. In addition, each of these sites modulates ARHGAP36 recruitment to the plasma membrane or primary cilium. Through comparative proteomics, we also have identified proteins that preferentially interact with active ARHGAP36, and we demonstrate that one binding partner, prolyl oligopeptidase-like protein, is a novel ARHGAP36 antagonist. Our work reveals multiple modes of ARHGAP36 regulation and establishes an experimental framework that can be applied towards other signaling proteins. This accession contains raw data from an interactome analysis of wild-type and L317P mutant ARHGAP36 isoform 2, represented in Fig. 5 and Dataset S2 of the associated publication.nactive GAP-like domain mutant, we have also identified prolyl oligopeptidase-like protein (PREPL) as a direct antagonist that decreases ARHGAP36 protein expression. In combination, our systems-level analyses uncover an ensemble of mechanisms that regulate ARHGAP36 expression, localization, and activity state, providing a potential means for context-specific ARHGAP36 functions. This accession contains raw data from an interactome analysis of wild-type and L317P mutant ARHGAP36 isoform 2, represented in Fig. 5 and Dataset S2 of the associated publication.

INSTRUMENT(S): Q Exactive HF-X, Orbitrap Fusion

ORGANISM(S): Mus Musculus (mouse)

SUBMITTER: Patricia Nano  

LAB HEAD: James K. Chen

PROVIDER: PXD019056 | Pride | 2021-05-18

REPOSITORIES: Pride