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[Preprint]. 2024 Jan 31:2022.08.23.22278845. [Version 3] doi: 10.1101/2022.08.23.22278845

Figure 4: DENND5A interacts with polarity proteins MUPP1 and PALS1.

Figure 4:

a, A recombinant GST-tagged peptide containing amino acids 700-720 of human DENND5A sequence was generated for use in pulldown experiments. The bolded residue corresponds to Arg710 that is affected in the cohort (R710H). b, Table indicating the number of peptides corresponding to MUPP1 and PALS1 found bound to each GST fusion peptide used in the pulldown/mass spectrometry experiment. c, Overexpressed human MUPP1- and PALS1-FLAG bind to GST-tagged DENND5A peptides. d, Residues 700-720 are shown in red in a space-fill model (left) and magnified view (right) of the predicted DENND5A protein structure from AlphaFold. Dotted lines indicate hydrogen bonds. e, The interface between the DENN and RUN1 domains of DENND5A comprise many charged residues. f, GST pulldown experiments show that FLAG-DENN and GST-RUN1/PLAT physically interact. g, Co-immunoprecipitations between GFP-DENND5A and MUPP1- and PALS1-FLAG show that DENND5A only binds the polarity proteins when the intramolecular DENN-RUN1 interaction is disrupted.