FRINK Linked Data Fragments server

Matches in SemOpenAlex for { <https://semopenalex.org/work/W2093202312> ?p ?o ?g. }

• W2093202312 endingPage "598" @default.
• W2093202312 startingPage "586" @default.
• W2093202312 abstract "Liprins are highly conserved scaffold proteins that regulate cell adhesion, cell migration, and synapse development by binding to diverse target proteins. The molecular basis governing liprin/target interactions is poorly understood. The liprin-α2/CASK complex structure solved here reveals that the three SAM domains of liprin-α form an integrated supramodule that binds to the CASK kinase-like domain. As supported by biochemical and cellular studies, the interaction between liprin-α and CASK is unique to vertebrates, implying that the liprin-α/CASK interaction is likely to regulate higher-order brain functions in mammals. Consistently, we demonstrate that three recently identified X-linked mental retardation mutants of CASK are defective in binding to liprin-α. We also solved the liprin-α/liprin-β SAM domain complex structure, which uncovers the mechanism underlying liprin heterodimerizaion. Finally, formation of the CASK/liprin-α/liprin-β ternary complex suggests that liprins can mediate assembly of target proteins into large protein complexes capable of regulating numerous cellular activities. Liprins are highly conserved scaffold proteins that regulate cell adhesion, cell migration, and synapse development by binding to diverse target proteins. The molecular basis governing liprin/target interactions is poorly understood. The liprin-α2/CASK complex structure solved here reveals that the three SAM domains of liprin-α form an integrated supramodule that binds to the CASK kinase-like domain. As supported by biochemical and cellular studies, the interaction between liprin-α and CASK is unique to vertebrates, implying that the liprin-α/CASK interaction is likely to regulate higher-order brain functions in mammals. Consistently, we demonstrate that three recently identified X-linked mental retardation mutants of CASK are defective in binding to liprin-α. We also solved the liprin-α/liprin-β SAM domain complex structure, which uncovers the mechanism underlying liprin heterodimerizaion. Finally, formation of the CASK/liprin-α/liprin-β ternary complex suggests that liprins can mediate assembly of target proteins into large protein complexes capable of regulating numerous cellular activities. The liprin-α2 SAM repeats/CASK kinase domain complex structure is solved A unique insertion between SAM1 and 2 is required for liprin-α2 to bind to CASK X-linked mental retardation mutants of CASK are defective in binding to liprin-α The liprin-α/β complex structure suggests a suprasignaling complex assembly mode Liprins, originally identified as binding partners of the receptor protein tyrosine phosphatase LAR (leukocyte common antigen-related) (Serra-Pagès et al., 1998Serra-Pagès C. Medley Q.G. Tang M. Hart A. Streuli M. Liprins, a family of LAR transmembrane protein-tyrosine phosphatase-interacting proteins.J. Biol. Chem. 1998; 273: 15611-15620Crossref PubMed Scopus (213) Google Scholar), are known to play roles in many cellular processes, including cell migration, cell adhesion, lymphatic vessel development, and synaptic development and activity (Astigarraga et al., 2010Astigarraga S. Hofmeyer K. Farajian R. Treisman J.E. Three Drosophila liprins interact to control synapse formation.J. Neurosci. 2010; 30: 15358-15368Crossref PubMed Scopus (51) Google Scholar, Choe et al., 2006Choe K.M. Prakash S. Bright A. Clandinin T.R. Liprin-alpha is required for photoreceptor target selection in Drosophila.Proc. Natl. Acad. Sci. USA. 2006; 103: 11601-11606Crossref PubMed Scopus (47) Google Scholar, Dai et al., 2006Dai Y. Taru H. Deken S.L. Grill B. Ackley B. Nonet M.L. Jin Y. SYD-2 Liprin-alpha organizes presynaptic active zone formation through ELKS.Nat. Neurosci. 2006; 9: 1479-1487Crossref PubMed Scopus (148) Google Scholar, Dunah et al., 2005Dunah A.W. Hueske E. Wyszynski M. Hoogenraad C.C. Jaworski J. Pak D.T. Simonetta A. Liu G. Sheng M. LAR receptor protein tyrosine phosphatases in the development and maintenance of excitatory synapses.Nat. Neurosci. 2005; 8: 458-467Crossref PubMed Scopus (191) Google Scholar, Kaufmann et al., 2002Kaufmann N. DeProto J. Ranjan R. Wan H. Van Vactor D. Drosophila liprin-alpha and the receptor phosphatase Dlar control synapse morphogenesis.Neuron. 2002; 34: 27-38Abstract Full Text Full Text PDF PubMed Scopus (224) Google Scholar, Olsen et al., 2005Olsen O. Moore K.A. Fukata M. Kazuta T. Trinidad J.C. Kauer F.W. Streuli M. Misawa H. Burlingame A.L. Nicoll R.A. Bredt D.S. Neurotransmitter release regulated by a MALS-liprin-alpha presynaptic complex.J. Cell Biol. 2005; 170: 1127-1134Crossref PubMed Scopus (96) Google Scholar, Schoch et al., 2002Schoch S. Castillo P.E. Jo T. Mukherjee K. Geppert M. Wang Y. Schmitz F. Malenka R.C. Südhof T.C. RIM1alpha forms a protein scaffold for regulating neurotransmitter release at the active zone.Nature. 2002; 415: 321-326Crossref PubMed Scopus (456) Google Scholar, Shen et al., 2007Shen J.C. Unoki M. Ythier D. Duperray A. Varticovski L. Kumamoto K. Pedeux R. Harris C.C. Inhibitor of growth 4 suppresses cell spreading and cell migration by interacting with a novel binding partner, liprin alpha1.Cancer Res. 2007; 67: 2552-2558Crossref PubMed Scopus (102) Google Scholar, Wyszynski et al., 2002Wyszynski M. Kim E. Dunah A.W. Passafaro M. Valtschanoff J.G. Serra-Pagès C. Streuli M. Weinberg R.J. Sheng M. Interaction between GRIP and liprin-alpha/SYD2 is required for AMPA receptor targeting.Neuron. 2002; 34: 39-52Abstract Full Text Full Text PDF PubMed Scopus (204) Google Scholar, Zhen and Jin, 1999Zhen M. Jin Y. The liprin protein SYD-2 regulates the differentiation of presynaptic termini in C. elegans.Nature. 1999; 401: 371-375Crossref PubMed Scopus (278) Google Scholar). Vertebrates contain two families of liprins, liprin-α and liprin-β, which have four (α1, α2, α3, and α4) and two (β1 and β2) members, respectively (Serra-Pagès et al., 1998Serra-Pagès C. Medley Q.G. Tang M. Hart A. Streuli M. Liprins, a family of LAR transmembrane protein-tyrosine phosphatase-interacting proteins.J. Biol. Chem. 1998; 273: 15611-15620Crossref PubMed Scopus (213) Google Scholar). In contrast, C. elegans and Drosophila each contain only one liprin-α, called Syd-2 and Dliprin-α, respectively (Kaufmann et al., 2002Kaufmann N. DeProto J. Ranjan R. Wan H. Van Vactor D. Drosophila liprin-alpha and the receptor phosphatase Dlar control synapse morphogenesis.Neuron. 2002; 34: 27-38Abstract Full Text Full Text PDF PubMed Scopus (224) Google Scholar, Zhen and Jin, 1999Zhen M. Jin Y. The liprin protein SYD-2 regulates the differentiation of presynaptic termini in C. elegans.Nature. 1999; 401: 371-375Crossref PubMed Scopus (278) Google Scholar). All isoforms of liprins share highly similar domain organizations, consisting of an N-terminal coiled-coil domain and a C-terminal liprin homology (LH) region comprised of three sterile alpha motif (SAM) domains (Kaufmann et al., 2002Kaufmann N. DeProto J. Ranjan R. Wan H. Van Vactor D. Drosophila liprin-alpha and the receptor phosphatase Dlar control synapse morphogenesis.Neuron. 2002; 34: 27-38Abstract Full Text Full Text PDF PubMed Scopus (224) Google Scholar, Serra-Pagès et al., 1998Serra-Pagès C. Medley Q.G. Tang M. Hart A. Streuli M. Liprins, a family of LAR transmembrane protein-tyrosine phosphatase-interacting proteins.J. Biol. Chem. 1998; 273: 15611-15620Crossref PubMed Scopus (213) Google Scholar, Zürner and Schoch, 2009Zürner M. Schoch S. The mouse and human Liprin-alpha family of scaffolding proteins: genomic organization, expression profiling and regulation by alternative splicing.Genomics. 2009; 93: 243-253Crossref PubMed Scopus (32) Google Scholar) (Figure 1A ). The N-terminal coiled coils of liprin-α act as binding regions for several synaptic proteins, including CAST, GIT1, RIM, and KIF1A (Ko et al., 2003aKo J. Kim S. Valtschanoff J.G. Shin H. Lee J.R. Sheng M. Premont R.T. Weinberg R.J. Kim E. Interaction between liprin-alpha and GIT1 is required for AMPA receptor targeting.J. Neurosci. 2003; 23: 1667-1677Crossref PubMed Google Scholar, Ko et al., 2003bKo J. Na M. Kim S. Lee J.R. Kim E. Interaction of the ERC family of RIM-binding proteins with the liprin-alpha family of multidomain proteins.J. Biol. Chem. 2003; 278: 42377-42385Crossref PubMed Scopus (122) Google Scholar, Schoch et al., 2002Schoch S. Castillo P.E. Jo T. Mukherjee K. Geppert M. Wang Y. Schmitz F. Malenka R.C. Südhof T.C. RIM1alpha forms a protein scaffold for regulating neurotransmitter release at the active zone.Nature. 2002; 415: 321-326Crossref PubMed Scopus (456) Google Scholar, Shin et al., 2003Shin H. Wyszynski M. Huh K.H. Valtschanoff J.G. Lee J.R. Ko J. Streuli M. Weinberg R.J. Sheng M. Kim E. Association of the kinesin motor KIF1A with the multimodular protein liprin-alpha.J. Biol. Chem. 2003; 278: 11393-11401Crossref PubMed Scopus (146) Google Scholar). The SAM repeats can bind to both phosphatases (e.g., LAR, PTPδ, and PTPσ) (Serra-Pagès et al., 1998Serra-Pagès C. Medley Q.G. Tang M. Hart A. Streuli M. Liprins, a family of LAR transmembrane protein-tyrosine phosphatase-interacting proteins.J. Biol. Chem. 1998; 273: 15611-15620Crossref PubMed Scopus (213) Google Scholar) and protein kinases (e.g., CASK) (Olsen et al., 2005Olsen O. Moore K.A. Fukata M. Kazuta T. Trinidad J.C. Kauer F.W. Streuli M. Misawa H. Burlingame A.L. Nicoll R.A. Bredt D.S. Neurotransmitter release regulated by a MALS-liprin-alpha presynaptic complex.J. Cell Biol. 2005; 170: 1127-1134Crossref PubMed Scopus (96) Google Scholar), although the molecular basis of these interactions is unknown. Additionally, the SAM repeats of liprin-α and liprin-β can form heterodimers (Serra-Pagès et al., 1998Serra-Pagès C. Medley Q.G. Tang M. Hart A. Streuli M. Liprins, a family of LAR transmembrane protein-tyrosine phosphatase-interacting proteins.J. Biol. Chem. 1998; 273: 15611-15620Crossref PubMed Scopus (213) Google Scholar), although again the underlying molecular mechanism is unknown. Much of our current understanding of the functions of liprin is derived from extensive studies in neurons. In C. elegans and Drosophila, loss-of-function mutations of liprin-α result in enlarged active zones, reduced synapse formation, impaired synaptic transmission, and mistargeting of axons (Choe et al., 2006Choe K.M. Prakash S. Bright A. Clandinin T.R. Liprin-alpha is required for photoreceptor target selection in Drosophila.Proc. Natl. Acad. Sci. USA. 2006; 103: 11601-11606Crossref PubMed Scopus (47) Google Scholar, Hofmeyer et al., 2006Hofmeyer K. Maurel-Zaffran C. Sink H. Treisman J.E. Liprin-alpha has LAR-independent functions in R7 photoreceptor axon targeting.Proc. Natl. Acad. Sci. USA. 2006; 103: 11595-11600Crossref PubMed Scopus (39) Google Scholar, Kaufmann et al., 2002Kaufmann N. DeProto J. Ranjan R. Wan H. Van Vactor D. Drosophila liprin-alpha and the receptor phosphatase Dlar control synapse morphogenesis.Neuron. 2002; 34: 27-38Abstract Full Text Full Text PDF PubMed Scopus (224) Google Scholar, Prakash et al., 2009Prakash S. McLendon H.M. Dubreuil C.I. Ghose A. Hwa J. Dennehy K.A. Tomalty K.M. Clark K.L. Van Vactor D. Clandinin T.R. Complex interactions amongst N-cadherin, DLAR, and Liprin-alpha regulate Drosophila photoreceptor axon targeting.Dev. Biol. 2009; 336: 10-19Crossref PubMed Scopus (33) Google Scholar, Zhen and Jin, 1999Zhen M. Jin Y. The liprin protein SYD-2 regulates the differentiation of presynaptic termini in C. elegans.Nature. 1999; 401: 371-375Crossref PubMed Scopus (278) Google Scholar), indicating that liprin-α plays critical roles in synaptic development and activity. The specific interaction between liprin-α and CASK in mammals hints that liprin-α may be directly involved in neurotransmitter release as well as the establishment of cell polarity via the highly conserved Veli (also called MALS or mLin-7)/CASK/Mint-1 (or X11α) tripartite complex (Biederer and Südhof, 2000Biederer T. Südhof T.C. Mints as adaptors. Direct binding to neurexins and recruitment of munc18.J. Biol. Chem. 2000; 275: 39803-39806Crossref PubMed Scopus (183) Google Scholar, Borg et al., 1999Borg J.P. Lõpez-Figueroa M.O. de Taddèo-Borg M. Kroon D.E. Turner R.S. Watson S.J. Margolis B. Molecular analysis of the X11-mLin-2/CASK complex in brain.J. Neurosci. 1999; 19: 1307-1316Crossref PubMed Google Scholar, Feng et al., 2004Feng W. Long J.F. Fan J.S. Suetake T. Zhang M. The tetrameric L27 domain complex as an organization platform for supramolecular assemblies.Nat. Struct. Mol. Biol. 2004; 11: 475-480Crossref PubMed Scopus (70) Google Scholar, Hata et al., 1996Hata Y. Butz S. Südhof T.C. CASK: a novel dlg/PSD95 homolog with an N-terminal calmodulin-dependent protein kinase domain identified by interaction with neurexins.J. Neurosci. 1996; 16: 2488-2494Crossref PubMed Google Scholar, Hsueh et al., 1998Hsueh Y.P. Yang F.C. Kharazia V. Naisbitt S. Cohen A.R. Weinberg R.J. Sheng M. Direct interaction of CASK/LIN-2 and syndecan heparan sulfate proteoglycan and their overlapping distribution in neuronal synapses.J. Cell Biol. 1998; 142: 139-151Crossref PubMed Scopus (277) Google Scholar, Maximov et al., 1999Maximov A. Südhof T.C. Bezprozvanny I. Association of neuronal calcium channels with modular adaptor proteins.J. Biol. Chem. 1999; 274: 24453-24456Crossref PubMed Scopus (252) Google Scholar, Kaech et al., 1998Kaech S.M. Whitfield C.W. Kim S.K. The LIN-2/LIN-7/LIN-10 complex mediates basolateral membrane localization of the C. elegans EGF receptor LET-23 in vulval epithelial cells.Cell. 1998; 94: 761-771Abstract Full Text Full Text PDF PubMed Scopus (303) Google Scholar, Olsen et al., 2005Olsen O. Moore K.A. Fukata M. Kazuta T. Trinidad J.C. Kauer F.W. Streuli M. Misawa H. Burlingame A.L. Nicoll R.A. Bredt D.S. Neurotransmitter release regulated by a MALS-liprin-alpha presynaptic complex.J. Cell Biol. 2005; 170: 1127-1134Crossref PubMed Scopus (96) Google Scholar, Olsen et al., 2007Olsen O. Funke L. Long J.F. Fukata M. Kazuta T. Trinidad J.C. Moore K.A. Misawa H. Welling P.A. Burlingame A.L. et al.Renal defects associated with improper polarization of the CRB and DLG polarity complexes in MALS-3 knockout mice.J. Cell Biol. 2007; 179: 151-164Crossref PubMed Scopus (35) Google Scholar, Samuels et al., 2007Samuels B.A. Hsueh Y.P. Shu T. Liang H. Tseng H.C. Hong C.J. Su S.C. Volker J. Neve R.L. Yue D.T. Tsai L.H. Cdk5 promotes synaptogenesis by regulating the subcellular distribution of the MAGUK family member CASK.Neuron. 2007; 56: 823-837Abstract Full Text Full Text PDF PubMed Scopus (90) Google Scholar, Schoch et al., 2002Schoch S. Castillo P.E. Jo T. Mukherjee K. Geppert M. Wang Y. Schmitz F. Malenka R.C. Südhof T.C. RIM1alpha forms a protein scaffold for regulating neurotransmitter release at the active zone.Nature. 2002; 415: 321-326Crossref PubMed Scopus (456) Google Scholar). Recently, several human genetic studies have revealed that CASK mutations are closely associated with mental retardations (Froyen et al., 2007Froyen G. Van Esch H. Bauters M. Hollanders K. Frints S.G. Vermeesch J.R. Devriendt K. Fryns J.P. Marynen P. Detection of genomic copy number changes in patients with idiopathic mental retardation by high-resolution X-array-CGH: important role for increased gene dosage of XLMR genes.Hum. Mutat. 2007; 28: 1034-1042Crossref PubMed Scopus (141) Google Scholar, Hayashi et al., 2008Hayashi S. Mizuno S. Migita O. Okuyama T. Makita Y. Hata A. Imoto I. Inazawa J. The CASK gene harbored in a deletion detected by array-CGH as a potential candidate for a gene causative of X-linked dominant mental retardation.Am. J. Med. Genet. A. 2008; 146A: 2145-2151Crossref PubMed Scopus (32) Google Scholar, Najm et al., 2008Najm J. Horn D. Wimplinger I. Golden J.A. Chizhikov V.V. Sudi J. Christian S.L. Ullmann R. Kuechler A. Haas C.A. et al.Mutations of CASK cause an X-linked brain malformation phenotype with microcephaly and hypoplasia of the brainstem and cerebellum.Nat. Genet. 2008; 40: 1065-1067Crossref PubMed Scopus (192) Google Scholar, Piluso et al., 2009Piluso G. D'Amico F. Saccone V. Bismuto E. Rotundo I.L. Di Domenico M. Aurino S. Schwartz C.E. Neri G. Nigro V. A missense mutation in CASK causes FG syndrome in an Italian family.Am. J. Hum. Genet. 2009; 84: 162-177Abstract Full Text Full Text PDF PubMed Scopus (61) Google Scholar, Tarpey et al., 2009Tarpey P.S. Smith R. Pleasance E. Whibley A. Edkins S. Hardy C. O'Meara S. Latimer C. Dicks E. Menzies A. et al.A systematic, large-scale resequencing screen of X-chromosome coding exons in mental retardation.Nat. Genet. 2009; 41: 535-543Crossref PubMed Scopus (428) Google Scholar), although the molecular basis for this association is unknown. The mutation-induced impairment of the liprin-α/CASK interaction may constitute the molecular mechanism underlying the involvement of CASK in mental retardation. In contrast to the direct physical and functional interaction between liprin-α and CASK observed in mammals, liprin-α does not seem to interact with the CASK ortholog in invertebrates. The loss of function of liprin-α/Syd-2 in C. elegans manifests itself in the form of defects in active zone assembly with enlarged zone areas (Zhen and Jin, 1999Zhen M. Jin Y. The liprin protein SYD-2 regulates the differentiation of presynaptic termini in C. elegans.Nature. 1999; 401: 371-375Crossref PubMed Scopus (278) Google Scholar), whereas the Lin-2 defective mutant displayed a vulvaless phenotype (Kaech et al., 1998Kaech S.M. Whitfield C.W. Kim S.K. The LIN-2/LIN-7/LIN-10 complex mediates basolateral membrane localization of the C. elegans EGF receptor LET-23 in vulval epithelial cells.Cell. 1998; 94: 761-771Abstract Full Text Full Text PDF PubMed Scopus (303) Google Scholar). It has also been demonstrated that, at least in HSN neurons in C. elegans, liprin-α/Syd-2 and Lin-2 neither colocalize nor genetically interact with each other (Dai et al., 2006Dai Y. Taru H. Deken S.L. Grill B. Ackley B. Nonet M.L. Jin Y. SYD-2 Liprin-alpha organizes presynaptic active zone formation through ELKS.Nat. Neurosci. 2006; 9: 1479-1487Crossref PubMed Scopus (148) Google Scholar, Patel et al., 2006Patel M.R. Lehrman E.K. Poon V.Y. Crump J.G. Zhen M. Bargmann C.I. Shen K. Hierarchical assembly of presynaptic components in defined C. elegans synapses.Nat. Neurosci. 2006; 9: 1488-1498Crossref PubMed Scopus (130) Google Scholar). Given the highly conserved nature of both proteins throughout evolution, it is conceptually challenging to explain the functional differences for liprin-α and CASK between mammals and lower eukaryotes. To understand the molecular basis of liprin-mediated protein complex assembly, we characterized the liprin-α/CASK and liprin-α/liprin-β interactions in detail. The crystal structure of the liprin-α SAM domain repeats in complex with the CaM kinase (CaMK) domain of CASK reveals unexpected binding modes for both SAM domains and the CaMK domain. Importantly, the structure of the liprin-α/CASK complex reveals that the liprin-α/CASK interaction is specific to vertebrates. We discovered that several mutants of CASK found in X-linked mental retardation (XLMR) patients have decreased affinities for liprin-α. Finally, the structure of the liprin-α/liprin-β SAM repeat complex provides mechanistic insights into the assembly of the CASK/liprin-α/liprin-β ternary suprascaffold. We chose liprin-α2 for our study because this isoform has been shown to interact specifically with CASK (Olsen et al., 2005Olsen O. Moore K.A. Fukata M. Kazuta T. Trinidad J.C. Kauer F.W. Streuli M. Misawa H. Burlingame A.L. Nicoll R.A. Bredt D.S. Neurotransmitter release regulated by a MALS-liprin-alpha presynaptic complex.J. Cell Biol. 2005; 170: 1127-1134Crossref PubMed Scopus (96) Google Scholar, Samuels et al., 2007Samuels B.A. Hsueh Y.P. Shu T. Liang H. Tseng H.C. Hong C.J. Su S.C. Volker J. Neve R.L. Yue D.T. Tsai L.H. Cdk5 promotes synaptogenesis by regulating the subcellular distribution of the MAGUK family member CASK.Neuron. 2007; 56: 823-837Abstract Full Text Full Text PDF PubMed Scopus (90) Google Scholar). The first SAM domain (SAM1) of liprin-α2 was shown to be necessary and sufficient for binding to CASK, and the CaMK domain and the first L27 domain of CASK were shown to be necessary for CASK to interact with liprin-α2 by yeast two-hybrid assay (Olsen et al., 2005Olsen O. Moore K.A. Fukata M. Kazuta T. Trinidad J.C. Kauer F.W. Streuli M. Misawa H. Burlingame A.L. Nicoll R.A. Bredt D.S. Neurotransmitter release regulated by a MALS-liprin-alpha presynaptic complex.J. Cell Biol. 2005; 170: 1127-1134Crossref PubMed Scopus (96) Google Scholar) (see Figure 1A for the domain organizations of the two proteins). We first tried to verify the above findings using purified proteins. We found that the inclusion of an ∼25-residue extension at both the N and C termini of the canonical SAM1 domain boundary (residues 866–1001 for the extended SAM1) is necessary to obtain folded SAM1 domain. To our surprise, a mixture of the extended SAM1 and CASK_CaMK (with and without the first L27 domain) was eluted as two separate peaks corresponding to the isolated CASK_CaMK and SAM1, respectively, in an analytical gel filtration column, indicating weak or no binding between these two proteins (data not shown). In contrast, the entire LH region of liprin-α2 (liprin-α2_LH), composed of its three SAM domains, coeluted with CASK_CaMK in the analytical gel filtration column with an elution volume indicative of a molecular mass of ∼70 kDa (i.e., a 1:1 liprin-α2_LH/CASK_CaMK complex) (Figure 1B). Isothermal titration calorimetry (ITC)-based assays showed that liprin-α2_LH binds to CASK_CaMK with a Kd of ∼0.6 μM (Figure 1C). Further extension of liprin-α2_LH did not enhance its CASK binding (Figure 1D), indicating that liprin-α2_LH contains the complete CASK binding sequence. Consistent with our gel filtration chromatography results, the extended SAM1 alone binds to CASK_CaMK with a much weaker affinity (Kd ∼9 μM) (Figures 1C, middle panel, and 1D). We further showed that the first L27 domain of CASK is not required for liprin-α2 binding, as CASK_CaMK alone displayed an affinity for liprin-α2 equal to that of CASK_CaMK together with the first L27 domain (Figure 1D). Taken together, the above biochemical data demonstrate that the three SAM domains of liprin-α2 and the CaMK domain of CASK constitute the necessary and sufficient elements for the two proteins to form a stable complex. The above finding also raises the intriguing question of how three SAM domains together recognize a protein kinase domain, although an isolated SAM domain has been shown to bind to the kinase domain of MAP kinases (Qiao et al., 2006Qiao F. Harada B. Song H. Whitelegge J. Courey A.J. Bowie J.U. Mae inhibits Pointed-P2 transcriptional activity by blocking its MAPK docking site.EMBO J. 2006; 25: 70-79Crossref PubMed Scopus (29) Google Scholar, Seidel and Graves, 2002Seidel J.J. Graves B.J. An ERK2 docking site in the Pointed domain distinguishes a subset of ETS transcription factors.Genes Dev. 2002; 16: 127-137Crossref PubMed Scopus (145) Google Scholar). To elucidate the molecular details of the liprin-α2/CASK interaction, we determined the crystal structure of the liprin-α2_LH/CASK_CaMK complex at 2.2 Å resolution (Table 1). Consistent with the data shown in Figure 1B, each asymmetric unit of the crystal contains one complex molecule with a 1:1 stoichiometry (Figure 1E). Except for a 20-residue flexible loop (aa 998–1017) connecting SAM1 and SAM2 of liprin-α2 and a few residues at the termini of the two proteins, the electron densities of the rest of the complex were clearly assigned.Table 1Statistics of Data Collection and Model RefinementData CollectionLiprin-α2_LH/CASK_CaMKLiprin-α2_LH/liprin-β1_LHSpace groupP41212P61Unit cell parameters (Å)a = b = 78.6, c = 227.4a = b = 141.9, c = 181.1Resolution range (Å)50–2.2 (2.24–2.2)50–2.9 (2.95–2.9)No. of unique reflections36535 (1780)45715 (2260)Redundancy5.9 (4.1)5.8 (5.2)I/σ20.4 (2.3)20.5 (2.2)Completeness (%)98.6 (96.6)99.5 (99.5)Rmerge (%)aRmerge = Σ|Ii − Im|/ΣIi, where Ii is the intensity of the measured reflection and Im is the mean intensity of all symmetry-related reflections.7.8 (55.4)6.8 (62.3)Structure RefinementResolution (Å)50–2.2 (2.26–2.2)50–2.9 (2.975–2.9)Rcryst/Rfree (%)bRcryst = Σ‖Fobs| − |Fcalc‖/Σ|Fobs|, where Fobs and Fcalc are observed and calculated structure factors. Rfree = ΣT‖Fobs| − |Fcalc‖/ΣT|Fobs|, where T is a test data set of about 5% of the total reflections randomly chosen and set aside prior to refinement.19.5 (24.3)/23.6 (31.5)21.4 (27.1)/25.8 (39.1)Rmsd bonds (Å)/angles (°)0.011/1.20.009/1.3Average B factor53.5102.4No. of atoms Protein atoms49398120 Water molecules11013 Other molecules3630No. of reflections Working set34,61942,766 Test set18182263Ramachandran plot Most favored regions (%)93.289.0 Additionally allowed (%)6.611.0 Generously allowed (%)0.20.0Numbers in parentheses represent the value for the highest-resolution shell.a Rmerge = Σ|Ii − Im|/ΣIi, where Ii is the intensity of the measured reflection and Im is the mean intensity of all symmetry-related reflections.b Rcryst = Σ‖Fobs| − |Fcalc‖/Σ|Fobs|, where Fobs and Fcalc are observed and calculated structure factors. Rfree = ΣT‖Fobs| − |Fcalc‖/ΣT|Fobs|, where T is a test data set of about 5% of the total reflections randomly chosen and set aside prior to refinement. Open table in a new tab Numbers in parentheses represent the value for the highest-resolution shell. In the complex (Figure 1E), CASK_CaMK adopts a typical protein kinase fold. The three SAM domains of liprin-α2 pack sequentially in a head-to-tail manner, forming a linear assembly. The interaction between CASK_CaMK and liprin-α2 is restricted to the C-lobe of the CaMK domain. The SAM1 and SAM2 domains of liprin-α2 make extensive contact with the bottom of the C-lobe of CaMK, and a unique insertion between SAM1 and SAM2 of liprin-α2 wraps around the backside of the CaMK C-lobe (relative to the CaMK catalytic cleft) (Figure 1E). The liprin-α2_LH/CASK_CaMK complex structure reveals a previously unknown interaction mode for SAM domains. SAM domains are best known to form homo- or heterodimers or oligomers and to interact with non-SAM protein domains and even RNAs (Qiao and Bowie, 2005Qiao F. Bowie J.U. The many faces of SAM.Sci. STKE. 2005; 2005: re7PubMed Google Scholar). The observation that three SAM domains in tandem are required for binding to a target protein is unprecedented. The structure of the liprin-α2_LH/CASK_CaMK complex also explains why the extended SAM1 (corresponding to the canonical SAM1 and the insertion between SAM1 and SAM2) is not sufficient for liprin-α2 to bind to CASK (Figures 1C and 1D). All liprins contain three highly conserved SAM domains arranged in tandem at their C-terminal ends (Figure 2A ). Although sequence similarities are low, the overall folds of the three SAM domains are similar (Figure S1A). In the liprin-α2 structure, the head-to-tail interactions between the three SAM domains lead to the formation of two interdomain interfaces (SAM1/2 and SAM2/3) (Figures 1E and 3A ). The head face of SAM1 and the tail side of SAM3 are open in the complex (Figures 1E); as we show below, these open sides mediate the formation of liprin-α/β heterodimers. The SAM domains in liprin-α2 are held together by H-bonding, hydrophobic, and charge-charge interactions. The interaction between SAM2 and SAM3 is further stabilized by a connecting α helix (αII-III), which physically staples the two domains together (Figure 2B). Consistent with the extensive interdomain interactions observed between SAM2 and 3, we were not able to obtain isolated SAM2 or SAM3 despite numerous trials with different domain boundaries and expression conditions. SAM1 contains an additional α helix (αN) at its N terminus; the extensive hydrophobic contact between αN and the canonical SAM1 explains why αN is essential for the preparation of stably folded SAM1 and SAM123 (Figure 2C). The interaction between SAM1 and SAM23 is strong, as we were able to obtain a highly stable SAM1/SAM23 complex simply by mixing isolated SAM1 with SAM23 (data not shown). The residues forming the inter-SAM interface in liprins are highly conserved (Figure 2A), suggesting that the linear head-to-tail assembly of the three SAM repeats is a feature common to all liprin isoforms. Since both SAM1 and SAM2 of liprin-α2 make direct contact with CASK_CaMK in the complex structure (Figures 1D and 1E), the formation of the structurally integrated SAM123 repeats is functionally required for the formation of the liprin-α2/CASK complex. Taken together, our structural and biochemical analysis reveals that liprin-α2 SAM123 acts as a structural and functional supramodule with its interaction mode that has not been previously described in SAM domains. Analysis of SAM domain proteins in mammalian genomes shows that a number of proteins contain multiple SAM domain repeats linked closely with each other in their primary sequences (Figure S1B). It is possible that the formation of SAM domain supramodules is a feature shared by many multi-SAM domain proteins. Indeed, a recent structural study of a tandem SAM domain protein called AIDA-1 showed that its two SAM domains indeed form a structural supramodule via a similar head-to-tail interaction mode (Kurabi et al., 2009Kurabi A. Brener S. Mobli M. Kwan J.J. Donaldson L.W. A Nuclear Localization Signal at the SAM-SAM Domain Interface of AIDA-1 Suggests a Requirement for Domain Uncoupling Prior to Nuclear Import.J. Mol. Biol. 2009; 392: 1168-1177Crossref PubMed Scopus (22) Google Scholar) (Figure 3A).Figure 3The Unique Assembly Mode of the SAM Repeats in Liprin-αShow full caption(A) Comparisons of the head-to-tail SAM/SAM interactions in liprin-α, in the polyhomeotic-SAM polymer (Ph-SAM, PDB code: 1KW4), and in the SAM tandem of AIDA-1 (PDB" @default.
• W2093202312 created "2016-06-24" @default.
• W2093202312 creator A5003353061 @default.
• W2093202312 creator A5021163747 @default.
• W2093202312 creator A5045266247 @default.
• W2093202312 creator A5058356091 @default.
• W2093202312 creator A5071825534 @default.
• W2093202312 creator A5089341631 @default.
• W2093202312 date "2011-08-01" @default.
• W2093202312 modified "2023-10-14" @default.
• W2093202312 title "Liprin-Mediated Large Signaling Complex Organization Revealed by the Liprin-α/CASK and Liprin-α/Liprin-β Complex Structures" @default.
• W2093202312 cites W1919311176 @default.
• W2093202312 cites W1964189494 @default.
• W2093202312 cites W1973160416 @default.
• W2093202312 cites W1978050645 @default.
• W2093202312 cites W1981434287 @default.
• W2093202312 cites W1991208570 @default.
• W2093202312 cites W1991626011 @default.
• W2093202312 cites W1999369847 @default.
• W2093202312 cites W2002025388 @default.
• W2093202312 cites W2005089183 @default.
• W2093202312 cites W2006384901 @default.
• W2093202312 cites W2014347276 @default.
• W2093202312 cites W2015889211 @default.
• W2093202312 cites W2017397242 @default.
• W2093202312 cites W2029815031 @default.
• W2093202312 cites W2031121891 @default.
• W2093202312 cites W2032244100 @default.
• W2093202312 cites W2041798139 @default.
• W2093202312 cites W2044935554 @default.
• W2093202312 cites W2045597434 @default.
• W2093202312 cites W2057347947 @default.
• W2093202312 cites W2075725530 @default.
• W2093202312 cites W2077576116 @default.
• W2093202312 cites W2080209385 @default.
• W2093202312 cites W2090378913 @default.
• W2093202312 cites W2093807978 @default.
• W2093202312 cites W2096865062 @default.
• W2093202312 cites W2097782660 @default.
• W2093202312 cites W2099387330 @default.
• W2093202312 cites W2112119382 @default.
• W2093202312 cites W2116921595 @default.
• W2093202312 cites W2118098860 @default.
• W2093202312 cites W2129082371 @default.
• W2093202312 cites W2130638580 @default.
• W2093202312 cites W2132369391 @default.
• W2093202312 cites W2133209165 @default.
• W2093202312 cites W2138301595 @default.
• W2093202312 cites W2140490140 @default.
• W2093202312 cites W2146624371 @default.
• W2093202312 cites W2152231606 @default.
• W2093202312 cites W2165554927 @default.
• W2093202312 cites W2169209199 @default.
• W2093202312 doi "https://doi.org/10.1016/j.molcel.2011.07.021" @default.
• W2093202312 hasPubMedId "https://pubmed.ncbi.nlm.nih.gov/21855798" @default.
• W2093202312 hasPublicationYear "2011" @default.
• W2093202312 type Work @default.
• W2093202312 sameAs 2093202312 @default.
• W2093202312 citedByCount "80" @default.
• W2093202312 countsByYear W20932023122012 @default.
• W2093202312 countsByYear W20932023122013 @default.
• W2093202312 countsByYear W20932023122014 @default.
• W2093202312 countsByYear W20932023122015 @default.
• W2093202312 countsByYear W20932023122016 @default.
• W2093202312 countsByYear W20932023122017 @default.
• W2093202312 countsByYear W20932023122018 @default.
• W2093202312 countsByYear W20932023122019 @default.
• W2093202312 countsByYear W20932023122020 @default.
• W2093202312 countsByYear W20932023122021 @default.
• W2093202312 countsByYear W20932023122022 @default.
• W2093202312 countsByYear W20932023122023 @default.
• W2093202312 crossrefType "journal-article" @default.
• W2093202312 hasAuthorship W2093202312A5003353061 @default.
• W2093202312 hasAuthorship W2093202312A5021163747 @default.
• W2093202312 hasAuthorship W2093202312A5045266247 @default.
• W2093202312 hasAuthorship W2093202312A5058356091 @default.
• W2093202312 hasAuthorship W2093202312A5071825534 @default.
• W2093202312 hasAuthorship W2093202312A5089341631 @default.
• W2093202312 hasBestOaLocation W20932023121 @default.
• W2093202312 hasConcept C2780858604 @default.
• W2093202312 hasConcept C53645450 @default.
• W2093202312 hasConcept C54355233 @default.
• W2093202312 hasConcept C62478195 @default.
• W2093202312 hasConcept C86803240 @default.
• W2093202312 hasConcept C95444343 @default.
• W2093202312 hasConceptScore W2093202312C2780858604 @default.
• W2093202312 hasConceptScore W2093202312C53645450 @default.
• W2093202312 hasConceptScore W2093202312C54355233 @default.
• W2093202312 hasConceptScore W2093202312C62478195 @default.
• W2093202312 hasConceptScore W2093202312C86803240 @default.
• W2093202312 hasConceptScore W2093202312C95444343 @default.
• W2093202312 hasIssue "4" @default.
• W2093202312 hasLocation W20932023121 @default.
• W2093202312 hasLocation W20932023122 @default.
• W2093202312 hasOpenAccess W2093202312 @default.
• W2093202312 hasPrimaryLocation W20932023121 @default.
• W2093202312 hasRelatedWork W1973329173 @default.
• W2093202312 hasRelatedWork W1977677498 @default.

• next