The Fe65 Adaptor Protein Interacts through Its PID1 Domain with the Transcription Factor CP2/LSF/LBP1

Abstract

The neural protein Fe65 possesses three putative protein-protein interaction domains: one WW domain and two phosphotyrosine interaction/phosphotyrosine binding domains (PID1 and PID2); the most C-terminal of these domains (PID2) interacts in vivo with the Alzheimer's β-amyloid precursor protein, whereas the WW domain binds to Mena, the mammalian homolog ofDrosophila-enabled protein. By the interaction trap procedure, we isolated a cDNA clone encoding a possible ligand of the N-terminal PID/PTB domain of Fe65 (PID1). Sequence analysis of this clone revealed that this ligand corresponded to the previously identified transcription factor CP2/LSF/LBP1. Co-immunoprecipitation experiments demonstrated that the interaction between Fe65 and CP2/LSF/LBP1 also takes place in vivo between the native molecules. The localization of both proteins was studied using fractionated cellular extracts. These experiments demonstrated that the various isoforms of CP2/LSF/LBP1 are differently distributed among subcellular fractions. At least one isoform, derived from alternative splicing (LSF-ID), is present outside the nucleus; Fe65 was found in both fractions. Furthermore, transfection experiments with an HA-tagged CP2/LSF/LBP1 cDNA demonstrated that Fe65 interacts also with the nuclear form of CP2/LSF/LBP1. Considering that the analysis of Fe65 distribution in fractionated cell extracts demonstrated that this protein is present both in nuclear and non-nuclear fractions, we examined the expression of Fe65 deletion mutants in the two fractions. This analysis allowed us to observe that a small region N-terminal to the WW domain is phosphorylated and is necessary for the presence of Fe65 in the nuclear fraction. The neural protein Fe65 possesses three putative protein-protein interaction domains: one WW domain and two phosphotyrosine interaction/phosphotyrosine binding domains (PID1 and PID2); the most C-terminal of these domains (PID2) interacts in vivo with the Alzheimer's β-amyloid precursor protein, whereas the WW domain binds to Mena, the mammalian homolog ofDrosophila-enabled protein. By the interaction trap procedure, we isolated a cDNA clone encoding a possible ligand of the N-terminal PID/PTB domain of Fe65 (PID1). Sequence analysis of this clone revealed that this ligand corresponded to the previously identified transcription factor CP2/LSF/LBP1. Co-immunoprecipitation experiments demonstrated that the interaction between Fe65 and CP2/LSF/LBP1 also takes place in vivo between the native molecules. The localization of both proteins was studied using fractionated cellular extracts. These experiments demonstrated that the various isoforms of CP2/LSF/LBP1 are differently distributed among subcellular fractions. At least one isoform, derived from alternative splicing (LSF-ID), is present outside the nucleus; Fe65 was found in both fractions. Furthermore, transfection experiments with an HA-tagged CP2/LSF/LBP1 cDNA demonstrated that Fe65 interacts also with the nuclear form of CP2/LSF/LBP1. Considering that the analysis of Fe65 distribution in fractionated cell extracts demonstrated that this protein is present both in nuclear and non-nuclear fractions, we examined the expression of Fe65 deletion mutants in the two fractions. This analysis allowed us to observe that a small region N-terminal to the WW domain is phosphorylated and is necessary for the presence of Fe65 in the nuclear fraction. Alzheimer's disease (AD) 1The abbreviations used are: ADAlzheimer's diseaseAββ-amyloid peptideAPPβ-amyloid precursor proteinCMVcytomegalovirus, CY, cytosolicHAhemagglutininPIDphosphotyrosine interaction domainPTBphosphotyrosine bindingPCRpolymerase chain reactionPAGEpolyacrylamide gel electrophoresis.1The abbreviations used are: ADAlzheimer's diseaseAββ-amyloid peptideAPPβ-amyloid precursor proteinCMVcytomegalovirus, CY, cytosolicHAhemagglutininPIDphosphotyrosine interaction domainPTBphosphotyrosine bindingPCRpolymerase chain reactionPAGEpolyacrylamide gel electrophoresis. is a neurodegenerative disorder in which the main pathological traits are neuronal loss and the presence of neurofibrillary tangles and senile plaques in affected tissues (1Selkoe D.J. J. Biol. Chem. 1996; 271: 18295-18298Abstract Full Text Full Text PDF PubMed Scopus (756) Google Scholar, 2Yanker B.A. Neuron. 1996; 16: 921-932Abstract Full Text Full Text PDF PubMed Scopus (912) Google Scholar). These plaques are formed by extracellular deposits of a 4-kDa peptide, known as β-amyloid (Aβ), which is derived from the proteolytic processing of the β-amyloid precursor protein (APP). APP is an integral membrane protein ubiquitously expressed at the level of plasma membrane and of several intracellular compartments (3Selkoe D.J. Annu. Rev. Cell Biol. 1994; 10: 373-403Crossref PubMed Scopus (740) Google Scholar). It shows a large extracellular/intraluminal domain, a single transmembrane tract, and a small cytosolic (CY) domain. Alzheimer's disease β-amyloid peptide β-amyloid precursor protein cytomegalovirus, CY, cytosolic hemagglutinin phosphotyrosine interaction domain phosphotyrosine binding polymerase chain reaction polyacrylamide gel electrophoresis. Alzheimer's disease β-amyloid peptide β-amyloid precursor protein cytomegalovirus, CY, cytosolic hemagglutinin phosphotyrosine interaction domain phosphotyrosine binding polymerase chain reaction polyacrylamide gel electrophoresis. Familial forms of AD are linked in a few cases to mutations of the APP gene, probably directly affecting the processing of APP and therefore Aβ generation. In other cases, mutations of presenilin 1 and 2 genes occur, which produce a phenotype that includes Aβ overproduction and deposition (4Haas C. Neuron. 1997; 18: 687-690Abstract Full Text Full Text PDF PubMed Scopus (143) Google Scholar, 5Hardy J. Trends Neurosci. 1997; 20: 154-159Abstract Full Text Full Text PDF PubMed Scopus (1267) Google Scholar) and, as suggested by recent results, could be a consequence of a direct interaction between APP and PS2 (6Xia W. Zhang J. Perez R. Koo E.H. Selkoe D.J. Proc. Natl. Acad. Sci. U. S. A. 1997; 94: 8208-8213Crossref PubMed Scopus (238) Google Scholar). Very little is known about APP and presenilin functions; consequently, our understanding of molecular mechanisms regulating APP processing is limited. The transmembrane topology of APP suggests that it could be involved in the transduction of signals through the membrane. Although the possible ligands of the extracellular/intraluminal domain of APP are still unknown, recent results showed that a complex protein network seems to be centered on the APP CY domain. In fact, at least four proteins have been demonstrated to interact with this 50-amino acid-long cytosolic tail. The first is the Go protein, which is activated bothin vitro and in vivo by direct interaction with APP (7Nishimoto I. Okamoto T. Matsuura Y. Takahashi S. Okamoto T. Murayama Y. Ogata E. Nature. 1993; 362: 75-79Crossref PubMed Scopus (371) Google Scholar). Mutant forms of APP constitutively activate Go (8Okamoto T. Takeda S. Murayama Y. Ogata E. Nishimoto I. J. Biol. Chem. 1995; 270: 4205-4208Abstract Full Text Full Text PDF PubMed Scopus (148) Google Scholar) and provoke various effects, including DNA fragmentation (9Giambarella U. Yamatsuji T. Okamoto T. Matsui T. Ikezu T. Murayama Y. Levine M.A. Katz A. Gautam N. Nishimoto I. EMBO J. 1997; 16: 4897-4907Crossref PubMed Scopus (82) Google Scholar). The second protein found to interact with APP was named APP-BP1. It is similar to the auxin resistance gene product AXR1 ofArabidopsis and to a protein in Caenorabditis elegans of unknown function (10Chow N. Korenberg J.R. Chen X.-N. Neve R.L. J. Biol. Chem. 1996; 271: 11339-11346Abstract Full Text Full Text PDF PubMed Scopus (118) Google Scholar). The X11 protein is neuron specific, and it interacts with APP through a phosphotyrosine interaction/phosphotyrosine binding (PID/PTB) domain and possesses other putative protein-protein interaction domains (11Borg J.P. Ooi J. Levy E. Margolis B. Mol. Cell. Biol. 1996; 16: 6229-6241Crossref PubMed Scopus (430) Google Scholar). The fourth member of this family of APP CY domain ligands is Fe65. It is a 90-kDa adaptor protein expressed in neurons of several regions of the mammalian nervous system (12Duilio A. Zambrano N. Mogavero A.R. Ammendola R. Cimino F. Russo T. Nucleic Acids Res. 1991; 19: 5269-5274Crossref PubMed Scopus (88) Google Scholar, 13Simeone A. Duilio A. Fiore F. Acampora D. De Felice C. Faraonio R. Paolocci F. Cimino F. Russo T. Dev. Neurosci. 1994; 16: 53-60Crossref PubMed Scopus (30) Google Scholar). It possesses three putative protein-protein interaction domains: one WW domain and two PID/PTB domains. We have demonstrated that the most C-terminal of these domains (PID2) binds both in vitro and in vivo to APP (14Fiore F. Zambrano N. Minopoli G. Donini V. Duilio A. Russo T. J. Biol. Chem. 1995; 270: 30853-30856Abstract Full Text Full Text PDF PubMed Scopus (267) Google Scholar, 15Zambrano N. Buxbaum J.D. Minopoli G. Fiore F. De Candia P. De Renzis S. Faraonio R. Sabo S. Cheetham J. Sudol M. Russo T. J. Biol. Chem. 1997; 272: 6399-6405Abstract Full Text Full Text PDF PubMed Scopus (139) Google Scholar). Furthermore, two other proteins similar to Fe65, named Fe65-L1 and Fe65-L2, were identified and found to interact with APP (16Guénette S.Y. Chen J. Jondro P.D. Tanzi R.E. Proc. Natl. Acad. Sci. U. S. A. 1996; 93: 10832-10837Crossref PubMed Scopus (149) Google Scholar, 17Duilio A. Faraonio R. Minopoli G. Zambrano N. Russo T. Biochem. J. 1998; 330: 513-519Crossref PubMed Scopus (86) Google Scholar). For interaction with Fe65 to occur, the last 32 C-terminal residues of APP are needed. These residues contain an NPXY motif, which was demonstrated to be the element of growth factor receptors recognized by PID/PTB domains of Shc and IRS1. However, contrary to observations of Shc and IRS1 behavior (18Blaikie P. Immanuel D. Wu J. Li N. Yajnic V. Margolis B. J. Biol. Chem. 1994; 269: 32031-32034Abstract Full Text PDF PubMed Google Scholar, 19Gustafson T.A. He W. Craparo A. Schaub C.D. O'Neil T.J. Mol. Cell. Biol. 1995; 15: 2500-2508Crossref PubMed Scopus (324) Google Scholar), the Fe65-APP interaction is phosphorylation independent (11Borg J.P. Ooi J. Levy E. Margolis B. Mol. Cell. Biol. 1996; 16: 6229-6241Crossref PubMed Scopus (430) Google Scholar, 15Zambrano N. Buxbaum J.D. Minopoli G. Fiore F. De Candia P. De Renzis S. Faraonio R. Sabo S. Cheetham J. Sudol M. Russo T. J. Biol. Chem. 1997; 272: 6399-6405Abstract Full Text Full Text PDF PubMed Scopus (139) Google Scholar). Mutations of APP responsible for familial forms of AD significantly affect the Fe65-APP in vivo interaction, especially in the case of the so-called Swedish mutant whose binding to Fe65 is almost completely abolished (15Zambrano N. Buxbaum J.D. Minopoli G. Fiore F. De Candia P. De Renzis S. Faraonio R. Sabo S. Cheetham J. Sudol M. Russo T. J. Biol. Chem. 1997; 272: 6399-6405Abstract Full Text Full Text PDF PubMed Scopus (139) Google Scholar). The possible role of the Fe65 adaptor protein was confirmed by the finding that several proteins interact with Fe65 through its WW domain. One of these ligands was identified as Mena, the mammalian homolog of the Drosophila enabled protein (20Ermekova K-S. Zambrano N. Linn H. Minopoli G. Gertler F. Russo T. Sudol M. J. Biol. Chem. 1997; 272: 32869-32874Abstract Full Text Full Text PDF PubMed Scopus (198) Google Scholar). To understand more about the function of Fe65, we addressed the point of the role of the third possible protein-protein interaction domain of Fe65, the N-terminal PID/PTB domain (PID1). Herein, we report that the PID1 domain of Fe65 interacts with the CP2/LSF/LBP1 protein, previously described as a transcription factor involved in the regulation of several genes (21Lim M. Mol. Cell. Biol. PubMed Google Scholar, D. U. Mol. Cell. Biol. 1994; PubMed Scopus Google Scholar, J. Biol. Chem. 1995; 270: Full Text Full Text PDF PubMed Scopus Google Scholar). of the of Fe65 and CP2/LSF/LBP1 the cell that these proteins are involved in a complex intracellular The system R. S. Proc. Natl. Acad. Sci. U. S. A. 1991; PubMed Scopus Google Scholar) was used to the Fe65 PID1 ligands and was as described in F. J. M. Cell and The region as a was from to of Fe65. This peptide the PID1 between and residues of are in the protein (16Guénette S.Y. Chen J. Jondro P.D. Tanzi R.E. Proc. Natl. Acad. Sci. U. S. A. 1996; 93: 10832-10837Crossref PubMed Scopus (149) Google Scholar). from to of the cDNA region R. Minopoli G. A. F. Cimino F. Russo T. Nucleic Acids Res. 1994; PubMed Scopus Google Scholar) was using a expression the The was used to the D. Nucleic Acids Res. 1994; PubMed Scopus Google Scholar) a clone that constitutively expressed the Fe65 region to the DNA binding domain of the transcription The the gene and the gene the of the transcription factor S. Nature. PubMed Scopus Google Scholar). The cDNA the which the was used to the were on in of and in at The were on the were these the was also as described in F. J. M. Cell and The from were and by The cDNA of the were by with and by with The various cDNA used in this were by of the cDNA previously described R. Minopoli G. A. F. Cimino F. Russo T. Nucleic Acids Res. 1994; PubMed Scopus Google the deletion mutant of Fe65 was by to 15Zambrano N. Buxbaum J.D. Minopoli G. Fiore F. De Candia P. De Renzis S. Faraonio R. Sabo S. Cheetham J. Sudol M. Russo T. J. Biol. Chem. 1997; 272: 6399-6405Abstract Full Text Full Text PDF PubMed Scopus (139) Google the of a mutant Fe65 cDNA an deletion of the region the WW and the PID1 domains to The and regions were by of cell using and DNA polymerase to the of the with the of The were by of the with and from with the gel in the of of for expression as a protein with a hemagglutinin The and the of the were by The and the were in with with and at in a were by at and with the For the expression (15Zambrano N. Buxbaum J.D. Minopoli G. Fiore F. De Candia P. De Renzis S. Faraonio R. Sabo S. Cheetham J. Sudol M. Russo T. J. Biol. Chem. 1997; 272: 6399-6405Abstract Full Text Full Text PDF PubMed Scopus (139) Google Scholar) was used in experiments in with the HA-tagged and expression with a expression the APP For the of the cellular were in and in 1 1 and of and The extracts were by at at and the protein was by the protein to extracts were to I. F. M. E. G. J. Biol. Chem. 1993; Full Text PDF PubMed Google 1 were in and in 1 1 of and were by at at for the was by at for at and used as the fraction. were in and on a by at for at The nuclear was by and in a 1 1 and of and The used in this been previously described (15Zambrano N. Buxbaum J.D. Minopoli G. Fiore F. De Candia P. De Renzis S. Faraonio R. Sabo S. Cheetham J. Sudol M. Russo T. J. Biol. Chem. 1997; 272: 6399-6405Abstract Full Text Full Text PDF PubMed Scopus (139) Google the Li G. Mol. Cell. Biol. 1994; PubMed Scopus Google Scholar) and its were a of R. and C. was from the was a of E. EMBO J. 1991; 10: PubMed Scopus Google Scholar), the was from and the was from The used for was a of Buxbaum of and the for the protein, was used as a For the the cellular extracts were with of the for 1 at and protein was to the extracts for of the The proteins were with a and by gel and to to the For the the were in in and with of the for 1 at The in was by of the in and a of protein was to the for 1 at were and the signals were by using the system For of protein from were with the the proteins were in and for at in the presence the of 1 of in a of the proteins were on and in experiments with the To for proteins to form with the region of Fe65 the PID1 motif, we the interaction trap The region of Fe65 from to including the PID1 element 15Zambrano N. Buxbaum J.D. Minopoli G. Fiore F. De Candia P. De Renzis S. Faraonio R. Sabo S. Cheetham J. Sudol M. Russo T. J. Biol. Chem. 1997; 272: 6399-6405Abstract Full Text Full Text PDF PubMed Scopus (139) Google Scholar), was the and used as a to a cDNA by the system in were the cDNA present in the were and the C-terminal of the known transcription factor CP2/LSF/LBP1 (21Lim M. Mol. Cell. Biol. PubMed Google Scholar, D. U. Mol. Cell. Biol. 1994; PubMed Scopus Google Scholar, J. Biol. Chem. 1995; 270: Full Text Full Text PDF PubMed Scopus Google Scholar). To the interaction in between of the two proteins also takes place in vivo between the native we the of in shows the of the proteins from cell extracts. The was with an with the and with the The of the by using the Fe65 with CP2/LSF/LBP1. The first the in this is the of in the CP2/LSF/LBP1 This element was found in of the proteins with PID/PTB both in the case of growth factor in which it be phosphorylated to interact with Shc IRS1 T.A. He W. Craparo A. Schaub C.D. O'Neil T.J. Mol. Cell. Biol. 1995; 15: 2500-2508Crossref PubMed Scopus (324) Google Scholar, K-S. Zambrano N. Linn H. Minopoli G. Gertler F. Russo T. Sudol M. J. Biol. Chem. 1997; 272: 32869-32874Abstract Full Text Full Text PDF PubMed Scopus (198) Google Scholar), and in the case of the of the APP whose interaction with the of the Fe65 family and with X11 on the phosphorylation independent (11Borg J.P. Ooi J. Levy E. Margolis B. Mol. Cell. Biol. 1996; 16: 6229-6241Crossref PubMed Scopus (430) Google Scholar, 15Zambrano N. Buxbaum J.D. Minopoli G. Fiore F. De Candia P. De Renzis S. Faraonio R. Sabo S. Cheetham J. Sudol M. Russo T. J. Biol. Chem. 1997; 272: 6399-6405Abstract Full Text Full Text PDF PubMed Scopus (139) Google Scholar). results have suggested that the is for Fe65-APP In fact, a acid-long peptide, the C-terminal of the APP and the motif, for the binding of Fe65, whereas an acid-long peptide a acid-long peptide, both the are to with APP for the interaction with Fe65 (15Zambrano N. Buxbaum J.D. Minopoli G. Fiore F. De Candia P. De Renzis S. Faraonio R. Sabo S. Cheetham J. Sudol M. Russo T. J. Biol. Chem. 1997; 272: 6399-6405Abstract Full Text Full Text PDF PubMed Scopus (139) Google Scholar). Furthermore, of the of APP demonstrated that the of of the C-terminal on the binding of Fe65 to APP (11Borg J.P. Ooi J. Levy E. Margolis B. Mol. Cell. Biol. 1996; 16: 6229-6241Crossref PubMed Scopus (430) Google Scholar). it seems that the Fe65 PID/PTB domains protein significantly from with the domains in Shc in IRS1 We examined the possible between the CY domains of APP family proteins and was The that the transcription factor whose intracellular localization is the forms with Fe65, as a of the cytosolic domain of the membrane protein APP and of the protein (20Ermekova K-S. Zambrano N. Linn H. Minopoli G. Gertler F. Russo T. Sudol M. J. Biol. Chem. 1997; 272: 32869-32874Abstract Full Text Full Text PDF PubMed Scopus (198) Google Scholar, M. J. P. Cell. 1996; Full Text Full Text PDF PubMed Scopus Google Scholar), is to by cytosolic with us to the subcellular of the two To we examined the presence of CP2/LSF/LBP1 in two cellular by and nuclear extracts were from as described and The of the nuclear was by to and the as this is to contain cytosolic proteins and the membrane proteins in the used for the cell the nuclear is to proteins present in To the of this we the distribution in the and of two the transcription factor A. Nucleic Acids Res. 1994; PubMed Scopus Google Scholar) and the cytosolic protein to the EMBO J. 1991; 10: PubMed Scopus Google Scholar, J. Biol. Chem. 1993; Full Text PDF PubMed Google Scholar). shows the with the and that is present in the nuclear and is present almost in the The extracts were by using the shows in the cell this recognized at least four protein between and The distribution of these among the two is In fact, the nuclear almost the four which have been described to to various phosphorylated forms of the protein De J. U. Dev. 1997; PubMed Google Scholar), whereas the the of whose is with that of the CP2/LSF/LBP1 by the D. U. Mol. Cell. Biol. 1994; PubMed Scopus Google Scholar). This CP2/LSF/LBP1 form is to the and therefore its presence outside the is To this we with a in which the the expression of the cDNA with the in the of fractionated cell extracts demonstrated that the is present in the and from the nuclear fraction. This suggests a role for the region from and present in the form of CP2/LSF/LBP1 in the subcellular distribution of the two for the of an complex that Fe65 interacts in the with the which is present outside the nuclear We directly this by cell extracts with the and the proteins with the of the of the CP2/LSF/LBP1 with the chain However, it is possible that the nuclear form of CP2/LSF/LBP1 interacts with Fe65. In fact, the distribution of Fe65 among the two fractions, in shows that most of the cellular Fe65 is in the a of Fe65 is also present in the nuclear fraction. we the interaction of Fe65 with the form of CP2/LSF/LBP1. To we with an expression in which an HA-tagged CP2/LSF/LBP1 cDNA is the of the shows that both in nuclear and in non-nuclear fractions, Fe65 with the HA-tagged CP2/LSF/LBP1. This that the Fe65 present in the nuclear is to form a complex with the form of CP2/LSF/LBP1. The presence of Fe65-APP was also found in both it be to the of a complex To this we the proteins with from with from with the encoding by with In both cases, was The of these results is that the complex the of this to this experiments are In a to be is that Fe65 and CP2/LSF/LBP1 form the with the nuclear independent from the demonstrated Fe65-APP and This be also observations which that Fe65 could function as a transcription factor (12Duilio A. Zambrano N. Mogavero A.R. Ammendola R. Cimino F. Russo T. Nucleic Acids Res. 1991; 19: 5269-5274Crossref PubMed Scopus (88) Google Scholar). point to be addressed with the molecular of the distribution of Fe65 between the nuclear and non-nuclear fractions. To this we various deletion mutants of the Fe65 described in that were in an expression the of the These were with an encoding the recognized by the The HA-tagged deletion mutants were expressed in by and expression was by using the in of the are expressed in and is in with the The most that be with this the of the In fact, as previously (15Zambrano N. Buxbaum J.D. Minopoli G. Fiore F. De Candia P. De Renzis S. Faraonio R. Sabo S. Cheetham J. Sudol M. Russo T. J. Biol. Chem. 1997; 272: 6399-6405Abstract Full Text Full Text PDF PubMed Scopus (139) Google Scholar), the Fe65 The be on the of the of isoforms of the Fe65 of similar proteins with the as the expression of the Fe65 cDNA in various cell to the of the it is that the is the of of the protein. Very among of the deletion the and proteins a of similar to that of the Fe65, whereas of the other are expressed as single This that the and proteins still contain the region of the still the for these possible that could at the level of the region of Fe65 present in the and in the two from the other is phosphorylation of the presence of several in the and in the that are known to be phosphorylated by and 1993; PubMed Scopus Google Scholar, PubMed Scopus Google Scholar). To this extracts from were with for various in the with results in the of the which the that Fe65 is To the possible role of the various Fe65 domains in the distribution of the proteins between the two described we the presence of the various deletion mutants in the two fractions. in the mutant protein is present in both fractions, the Fe65 and the protein whereas the mutants are significantly present in the fraction. This suggests that the presence of Fe65 in the nuclear is This to be on the of Fe65 with through its binding to the mutant the which is to the be with APP Furthermore, it is to that the of of and mutant Fe65 in the nuclear is the and that the phosphorylated forms of Fe65 are with the nuclear fraction. To the role of the small region the WW domain at its N-terminal in which the phosphorylated residues are probably we a deletion mutant of the Fe65 similar to the for an deletion the residues present between the WW and the PID1 domains The protein by this mutant is expressed in at and is present in the therefore that the small region the WW domain at its N-terminal is necessary to Fe65 to the nuclear fraction. The analysis of the three of the Fe65 protein family shows that Fe65 and Fe65-L1 a N-terminal the WW domain, whereas in it is significantly that its putative is residues of the WW domain A. Faraonio R. Minopoli G. Zambrano N. Russo T. Biochem. J. 1998; 330: 513-519Crossref PubMed Scopus (86) Google Scholar). The between Fe65 and Fe65-L1 is in regions the three protein-protein interaction and, the of the two proteins are for a small of that is in the region of Fe65 and that seems to be responsible for the of the protein This region is in finding the that this region of Fe65 The analysis of its that the most that could be involved in this regulation is by the present in the N-terminal of Fe65 the WW domain. These were found in DNA binding proteins and were suggested to be a in DNA binding M. J. Mol. Biol. PubMed Scopus Google Scholar). In CP2/LSF/LBP1 several of these are present and phosphorylated by protein J. U. Dev. 1997; PubMed Scopus Google Scholar). are also found in the an integral protein of the nuclear membrane C.D. G. J. Cell Biol. PubMed Scopus Google Scholar), and in the protein, also found in the nuclear membrane M. M.A. C. D. B. J. Cell Biol. 1993; PubMed Scopus Google Scholar). both have been demonstrated to be present at the level of the nuclear membrane J. M. H. J. H. Cell. 1997; Full Text Full Text PDF PubMed Scopus Google Scholar), and both contain several J. J. H. Proc. Natl. Acad. Sci. U. S. A. 1995; PubMed Scopus Google Scholar). Furthermore, APP is phosphorylated by at which is by a T. M. P. EMBO J. 1994; PubMed Scopus Google Scholar). We R. and C. for the E. for the and M. Sudol and F. Gertler for