Abstract
Several regulated mRNAs were detected by applying differential display to the mouse cerebellum during postnatal development. One cDNA fragment, referred to as CPD1 (GenBank U89345), was characterized and cloned. Northern blots showed maximum mRNA expression at postnatal day seven (P7). The mRNA encodes a protein of 260 amino acids. In situ RT-PCR showed that CPD1 is expressed mainly in granule cells and faintly in Purkinje cells. Polyclonal rabbit antibodies and oligobodies (oligonucleotide-based synthetic antibodies) revealed a protein of 34 kDa in Western blots. Immunohistochemistry showed not only marked nuclear staining but also mild cytoplasmic localization. Granule cells undergoing active division (P4) showed very little expression of CPD1 protein, which increases from P7 to P17. CPD1, affinity-purified using a chemically synthesized oligobody inhibits the activity of protein phosphatase PP2A but not protein phosphatase PP1. Differentiated PC12 cells also showed nuclear and cytoplasmic localization. Interestingly, maximal cytoplasmic CPD1/PP2A colocalization was observed near cell membrane regions that are far from growing neurites, and on growing cones. These results suggest that CPD1 might have an important role in cerebellar development.
Publication types
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Comparative Study
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Research Support, Non-U.S. Gov't
MeSH terms
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Amino Acid Sequence
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Animals
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Atrial Natriuretic Factor
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Blotting, Northern
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Blotting, Western
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Cell Division
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Cerebellar Cortex / growth & development
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Cerebellar Cortex / metabolism*
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DNA Primers
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DNA, Complementary / genetics
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Enzyme Inhibitors / chemistry
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Gene Expression Profiling*
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Gene Expression Regulation, Developmental*
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Gene Expression Regulation, Neoplastic
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Intracellular Signaling Peptides and Proteins
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Mice
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Mice, Inbred C57BL
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Molecular Chaperones
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Molecular Sequence Data
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Molecular Weight
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Morphogenesis
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Nerve Tissue Proteins / biosynthesis*
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Nerve Tissue Proteins / chemistry
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Nerve Tissue Proteins / genetics*
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Nuclear Proteins
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PC12 Cells / metabolism
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Peptide Fragments
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Phosphoprotein Phosphatases / antagonists & inhibitors
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Protein Precursors
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Proteins / chemistry
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Purkinje Cells / metabolism
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RNA, Messenger / biosynthesis
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RNA, Neoplasm / biosynthesis
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RNA-Binding Proteins
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Rats
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Reverse Transcriptase Polymerase Chain Reaction
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Sequence Alignment
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Sequence Homology, Amino Acid
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Synapses / physiology
Substances
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ANP32A protein, human
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Anp32e protein, mouse
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DNA Primers
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DNA, Complementary
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Enzyme Inhibitors
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Intracellular Signaling Peptides and Proteins
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Molecular Chaperones
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Nerve Tissue Proteins
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Nuclear Proteins
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Peptide Fragments
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Protein Precursors
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Proteins
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RNA, Messenger
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RNA, Neoplasm
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RNA-Binding Proteins
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atrial natriuretic factor prohormone (1-30), human
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Atrial Natriuretic Factor
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Phosphoprotein Phosphatases