The Fe65 adaptor protein interacts through its PID1 domain with the transcription factor CP2/LSF/LBP1.
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 beta-amyloid precursor protein, whereas the WW domain binds to Mena, the mammalian homolog of Drosophila-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.
Mesh Terms:
Amino Acid Sequence, Animals, Binding Sites, Carrier Proteins, Cell Fractionation, Cell Line, Cloning, Molecular, Cytoskeletal Proteins, DNA-Binding Proteins, Molecular Sequence Data, Nerve Tissue Proteins, Nuclear Proteins, Phosphorylation, RNA-Binding Proteins, Rats, Recombinant Proteins, Sequence Alignment, Sequence Analysis, DNA, Sequence Deletion, Transcription Factors, Transfection
Amino Acid Sequence, Animals, Binding Sites, Carrier Proteins, Cell Fractionation, Cell Line, Cloning, Molecular, Cytoskeletal Proteins, DNA-Binding Proteins, Molecular Sequence Data, Nerve Tissue Proteins, Nuclear Proteins, Phosphorylation, RNA-Binding Proteins, Rats, Recombinant Proteins, Sequence Alignment, Sequence Analysis, DNA, Sequence Deletion, Transcription Factors, Transfection
J. Biol. Chem.
Date: Aug. 07, 1998
PubMed ID: 9685356
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