title: Mutually exclusive binding of APPL(PH) to BAR domain and Reptin regulates β-catenin dependent transcriptional events.
abstract: Reptin functions in a wide range of biological processes including chromatin remodelling, nucleolar organization and transcriptional regulation of WNT signalling. As β-catenin dependent transcriptional repression and activation events involve binding of Reptin and histone deacetylase 1 to APPL endocytic proteins, this complex has become an important target to identify molecules governing endocytic processes and WNT signalling. Here, we describe the structural basis of APPL binding to Reptin to explore their mode of binding in context with APPL1/APPL2 dimerization. There is an evidence that both PH and BAR domains of APPL proteins exhibit alternately conserved regions involved in hetero-dimerization process and our in-silico data also corroborate this fact. Moreover, APPL2(PH) domain binds to the BAR domain region encompassing a nuclear localization signal. We conclude that APPL(PH) binding to BAR domain and Reptin is mutually exclusive which regulates the nucleocytoplasmic shuttling of Reptin. Furthermore, Reptin is unable to bind with membrane-associated APPL proteins. These observations were further expanded by experimental approaches where we identified a novel point mutation D316N lying in the APPL1(PH) domain which resulted in a significantly reduced binding with Reptin. By luciferase assays, we observed that overexpression of APPL1(D316N) and APPL1(WT) stimulated β-catenin/TCF dependent transcriptional activity in a similar manner which suggested that binding of Reptin to APPL1 is not necessary for β-catenin dependent target gene expression. Overall, our data attempt to highlight a comparative role of APPL proteins in controlling β-catenin dependent transcription mechanism which may improve our understanding of gene regulation.
authors.affiliation: National Center for Bioinformatics, Quaid-i-Azam University, Islamabad, Pakistan. Electronic address: sajidrwp@yahoo.co.uk.
authors.forename: Sajid
authors.initials: S
authors.lastname: Rashid
authors.forename: Zahida
authors.initials: Z
authors.lastname: Parveen
authors.forename: Saba
authors.initials: S
authors.lastname: Ferdous
authors.forename: Nousheen
authors.initials: N
authors.lastname: Bibi
mesh_terms: D000074183:ATPases Associated with Diverse Cellular Activities; D048868:Adaptor Proteins, Signal Transducing / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D000465:Algorithms; D000595:Amino Acid Sequence; D001665:Binding Sites; D002352:Carrier Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D004265:DNA Helicases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D057809:HEK293 Cells; D006801:Humans; D008958:Models, Molecular; D008969:Molecular Sequence Data; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D016415:Sequence Alignment; D014158:Transcription, Genetic; D015533:Transcriptional Activation / Q000235:genetics; D051176:beta Catenin / Q000378:metabolism*
chemical_list: C464801:APPL1 protein, human; C528555:APPL2 protein, human; D048868:Adaptor Proteins, Signal Transducing; D002352:Carrier Proteins; D051176:beta Catenin; D000074183:ATPases Associated with Diverse Cellular Activities; D004265:DNA Helicases; C122079:RUVBL2 protein, human
keywords: APPL; BAR domain; Molecular docking; PH domain; Reptin; β-Catenin