title: SHOC2 subcellular shuttling requires the KEKE motif-rich region and N-terminal leucine-rich repeat domain and impacts on ERK signalling.
abstract: SHOC2 is a scaffold protein composed almost entirely by leucine-rich repeats (LRRs) and having an N-terminal region enriched in alternating lysine and glutamate/aspartate residues (KEKE motifs). SHOC2 acts as a positive modulator of the RAS-RAF-MEK-ERK signalling cascade by favouring stable RAF1 interaction with RAS. We previously reported that the p.Ser2Gly substitution in SHOC2 underlies Mazzanti syndrome, a RASopathy clinically overlapping Noonan syndrome, promoting N-myristoylation and constitutive targeting of the mutant to the plasma membrane. We also documented transient nuclear translocation of wild-type SHOC2 upon EGF stimulation, suggesting a more complex function in signal transduction.Here, we characterized the domains controlling SHOC2 shuttling between the nucleus and cytoplasm, and those contributing to SHOC2S2Gmistargeting to the plasma membrane, analysed the structural organization of SHOC2's LRR motifs, and determined the impact of SHOC2 mislocalization on ERK signalling. We show that LRRs 1 to 13 constitute a structurally recognizable domain required for SHOC2 import into the nucleus and constitutive targeting of SHOC2S2Gto the plasma membrane, while the KEKE motif-rich region is necessary to achieve efficient SHOC2 export from the nucleus. We also document that SHOC2S2Glocalizes both in raft and non-raft domains, and that it translocates to the non-raft domains following stimulation. Finally, we demonstrate that SHOC2 trapping at different subcellular sites has a diverse impact on ERK signalling strength and dynamics, suggesting a dual counteracting modulatory role of SHOC2 in the control of ERK signalling exerted at different intracellular compartments.
authors.affiliation: Genetics and Rare Diseases Research Division, Ospedale Pediatrico Bambino Gesù, Rome, Italy.
authors.forename: Marialetizia
authors.initials: M
authors.lastname: Motta
authors.affiliation: CINECA, SCAI-SuperComputing Applications and Innovation Department, Rome, Italy.
authors.forename: Giovanni
authors.initials: G
authors.lastname: Chillemi
authors.affiliation: Department of Hematology, Oncology and Molecular Medicine.
authors.forename: Valentina
authors.initials: V
authors.lastname: Fodale
authors.affiliation: Department of Cell Biology and Neurosciences.
authors.forename: Serena
authors.initials: S
authors.lastname: Cecchetti
authors.affiliation: Italian National Centre for Rare Diseases, Istituto Superiore di Sanità, Rome, Italy.
authors.forename: Simona
authors.initials: S
authors.lastname: Coppola
authors.affiliation: Department of Hematology, Oncology and Molecular Medicine.
authors.forename: Silvia
authors.initials: S
authors.lastname: Stipo
authors.affiliation: Department of Hematology, Oncology and Molecular Medicine.
authors.forename: Viviana
authors.initials: V
authors.lastname: Cordeddu
authors.affiliation: Department of Cell Biology and Neurosciences.
authors.forename: Pompeo
authors.initials: P
authors.lastname: Macioce
authors.affiliation: The Mindich Child Health and Development Institute, Icahn School of Medicine at Mount Sinai, New York, NY, USA.
authors.forename: Bruce D
authors.initials: BD
authors.lastname: Gelb
authors.affiliation: Genetics and Rare Diseases Research Division, Ospedale Pediatrico Bambino Gesù, Rome, Italy marco.tartaglia@opbg.net.
authors.forename: Marco
authors.initials: M
authors.lastname: Tartaglia
mesh_terms: D020816:Amino Acid Motifs; D000818:Animals; D019556:COS Cells; D002460:Cell Line; D002467:Cell Nucleus / Q000378:metabolism*; D002522:Chlorocebus aethiops; D003593:Cytoplasm / Q000378:metabolism*; D057809:HEK293 Cells; D006801:Humans; D047908:Intracellular Signaling Peptides and Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D058247:Loose Anagen Hair Syndrome / Q000235:genetics*; D020935:MAP Kinase Signaling System; D051379:Mice; D041681:NIH 3T3 Cells; D009634:Noonan Syndrome / Q000235:genetics*; D021381:Protein Transport
chemical_list: D047908:Intracellular Signaling Peptides and Proteins; C112913:SHOC2 protein, human; C000622015:Shoc2 protein, mouse