ABSTRACT
Paraneoplastic neurological disorders may result from autoimmunity directed against antigens shared by the affected neurons and the associated cancer cells. We have recently reported the case of a woman with breast cancer and paraneoplastic lower motor neuron syndrome whose serum contained autoantibodies directed against axon initial segments and nodes of Ranvier of myelinated axons, including the axons of motoneurons. Here, we show that major targets of the autoantibodies of this patient are betaIVSigma1 spectrin and betaIV spectrin 140, two isoforms of the novel betaIV spectrin gene, as well as a neuronal surface epitope yet to be identified. Partial improvement of the neurological symptoms following cancer removal was associated with a drastic reduction in the titer of the autoantibodies against betaIV spectrin and nodal antigens in general, consistent with the autoimmune pathogenesis of the paraneoplastic lower motor neuron syndrome. The identification of betaIV spectrin isoforms and surface nodal antigens as novel autoimmune targets in lower motor neuron syndrome provide new insights into the pathogenesis of this severe neurological disease.
Subject(s)
Autoimmunity , Motor Neuron Disease/immunology , Nerve Tissue Proteins/immunology , Paraneoplastic Syndromes/immunology , Spectrin/immunology , Autoantibodies/analysis , Breast Neoplasms/immunology , Female , G(M1) Ganglioside/immunology , Humans , Molecular WeightABSTRACT
We report the identification of betaIV spectrin, a novel spectrin isolated as an interactor of the receptor tyrosine phosphatase-like protein ICA512. The betaIV spectrin gene is located on human and mouse chromosomes 19q13.13 and 7b2, respectively. Alternative splicing of betaIV spectrin generates at least four distinct isoforms, numbered betaIVSigma1-betaIVSigma4 spectrin. The longest isoform (betaIVSigma1 spectrin) includes an actin-binding domain, followed by 17 spectrin repeats, a specific domain in which the amino acid sequence ERQES is repeated four times, several putative SH3-binding sites and a pleckstrin homology domain. betaIVSigma2 and betaIVSigma3 spectrin encompass the NH(2)- and COOH-terminal halves of betaIVSigma1 spectrin, respectively, while betaIVSigma4 spectrin lacks the ERQES and the pleckstrin homology domain. Northern blots revealed an abundant expression of betaIV spectrin transcripts in brain and pancreatic islets. By immunoblotting, betaIVSigma1 spectrin is recognized as a protein of 250 kD. Anti-betaIV spectrin antibodies also react with two additional isoforms of 160 and 140 kD. These isoforms differ from betaIVSigma1 spectrin in terms of their distribution on subcellular fractionation, detergent extractability, and phosphorylation. In islets, the immunoreactivity for betaIV spectrin is more prominent in alpha than in beta cells. In brain, betaIV spectrin is enriched in myelinated neurons, where it colocalizes with ankyrin(G) 480/270-kD at axon initial segments and nodes of Ranvier. Likewise, betaIV spectrin is concentrated at the nodes of Ranvier in the rat sciatic nerve. In the rat hippocampus, betaIVSigma1 spectrin is detectable from embryonic day 19, concomitantly with the appearance of immunoreactivity at the initial segments. Thus, we suggest that betaIVSigma1 spectrin interacts with ankyrin(G) 480/270-kD and participates in the clustering of voltage-gated Na(+) channels and cell-adhesion molecules at initial segments and nodes of Ranvier.
Subject(s)
Axons/chemistry , Brain Chemistry/physiology , Nerve Tissue Proteins/analysis , Nerve Tissue Proteins/genetics , Ranvier's Nodes/chemistry , Sciatic Nerve/chemistry , Spectrin/analysis , Spectrin/genetics , Amino Acid Sequence , Animals , Ankyrins/metabolism , Autoantigens , Axons/physiology , Blood Proteins/chemistry , Blood Proteins/genetics , COS Cells , Chromosomes , Cloning, Molecular , Cytoplasm/chemistry , Cytoplasm/metabolism , Cytoskeleton/chemistry , Cytoskeleton/metabolism , Diabetic Neuropathies/physiopathology , Gene Expression/physiology , Hippocampus/chemistry , Hippocampus/cytology , Hippocampus/physiology , Humans , Islets of Langerhans/chemistry , Islets of Langerhans/physiology , Male , Membrane Proteins/analysis , Membrane Proteins/metabolism , Mice , Molecular Sequence Data , Nerve Tissue Proteins/chemistry , Phosphoproteins/chemistry , Phosphoproteins/genetics , Protein Structure, Tertiary , Protein Tyrosine Phosphatases/analysis , Protein Tyrosine Phosphatases/metabolism , RNA, Messenger/analysis , Ranvier's Nodes/physiology , Rats , Rats, Sprague-Dawley , Receptor-Like Protein Tyrosine Phosphatases, Class 8 , Sciatic Nerve/cytology , Sciatic Nerve/physiology , Signal Transduction/physiology , Sodium Channels/metabolism , Spectrin/chemistryABSTRACT
Islet cell autoantigen (ICA) 512 of type I diabetes is a receptor tyrosine phosphatase-like protein associated with the secretory granules of neurons and endocrine cells including insulin-secreting beta-cells of the pancreas. Here we show that in a yeast two-hybrid assay its cytoplasmic domain binds beta2-syntrophin, a modular adapter which in muscle cells interacts with members of the dystrophin family including utrophin, as well as the signaling molecule neuronal nitric oxide synthase (nNOS). The cDNA isolated by two-hybrid screening corresponded to a novel beta2-syntrophin isoform with a predicted molecular mass of 28 kDa. This isoform included the PDZ domain, but not the C-terminal region, which in full-length beta2-syntrophin is responsible for binding dystrophin-related proteins. In vitro binding of the beta2-syntrophin PDZ domain to ICA512 required both ICA512's C-terminal region and an internal polypeptide preceding its tyrosine phosphatase-like domain. Immunomicroscopy and co-immunoprecipitations from insulinoma INS-1 cells confirmed the occurrence of ICA512-beta2-syntrophin complexes in vivo. ICA512 also interacted in vitro with the PDZ domain of nNOS and ICA512-nNOS complexes were co-immunoprecipitated from INS-1 cells. Finally, we show that INS-1 cells, like muscle cells, contain beta2-syntrophin-utrophin oligomers. Thus, we propose that ICA512, through beta2-syntrophin and nNOS, links secretory granules with the actin cytoskeleton and signaling pathways involving nitric oxide.