Molecular cloning of a 2',3'-cyclic nucleotide 3'-phosphodiesterase: mRNAs with different 5' ends encode the same set of proteins in nervous and lymphoid tissues

Bernier, Lise; Álvarez, Fernando; Em, Norgard; Raible, David W.; Mentaberry, Alejandro; Jg, Schembri; Dd, Sabatini; Dr, Colman

doi:10.1523/jneurosci.07-09-02703.1987

Cited by 137 publications

(100 citation statements)

References 38 publications

(33 reference statements)

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“…Plasmid Description-All recombinant CNP expression vectors were generated from the rat CNP1 and CNP2 cDNA clones (14,32). First, a BamHI site, 17 nucleotides upstream of the ATG start codon of CNP1, was previously engineered by polymerase chain reaction to generate the vector, pBS/CNP1 (12).…”

Section: Methodsmentioning

confidence: 99%

Identification of Essential Residues in 2′,3′-Cyclic Nucleotide 3′-Phosphodiesterase

Lee¹,

Gravel²,

Gao

et al. 2001

Journal of Biological Chemistry

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2,3-Cyclic nucleotide 3-phosphodiesterase (CNP; EC 3.1.4.37) catalyzes in vitro hydrolysis of 3-phosphodiester bonds in 2,3-cyclic nucleotides to produce 2-nucleotides exclusively. N-terminal deletion mapping of the C-terminal two-thirds of recombinant rat CNP1 identified a region that possesses the catalytic domain, with further truncations abolishing activity. Proteolysis and kinetic analysis indicated that this domain forms a compact globular structure and contains all of the catalytically essential features. Subsequently, this catalytic fragment of CNP1 (CNP-CF) was used for chemical modification studies to identify amino acid residues essential for activity. 5,5-Dithiobis-(2-nitrobenzoic acid) modification studies and kinetic analysis of cysteine CNP-CF mutants revealed the nonessential role of cysteines for enzymatic activity. On the other hand, modification studies with diethyl pyrocarbonate indicated that two histidines are essential for CNPase activity. Consequently, the only two conserved histidines, His-230 and His-309, were mutated to phenylalanine and leucine. All four histidine mutants had k cat values 1000-fold lower than wild-type CNP-CF, but K m values were similar. Circular dichroism studies demonstrated that the low catalytic activities of the histidine mutants were not due to gross changes in secondary structure. Taken together, these results demonstrate that both histidines assume critical roles for catalysis.

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Section: Methodsmentioning

confidence: 99%

Identification of Essential Residues in 2′,3′-Cyclic Nucleotide 3′-Phosphodiesterase

Lee¹,

Gravel²,

Gao

et al. 2001

Journal of Biological Chemistry

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“…The blots were then UV-cross-linked (0.12 J), prehybridized, hybridized, and washed using standard techniques (Sambrook et al, 1989). The following probes were used: a f ull-length 1.25 kb Gtx cDNA, a 0.85 kb Pst fragment from a rat C N P cDNA (Bernier et al, 1987), a 1.5 kb EcoRI fragment from a rat M BP cDNA isolated in our laboratory, a 1.4 kb fragment from a rat PL P cDNA , and a 1.3 kb cDNA encoding rat glyceraldehyde 3-phosphate dehydrogenase (GAPDH) (Fort et al, 1985). 32 P-labeled probes were prepared by primer extension with random hexamers using the Prime-a-gene kit (Promega, Madison, W I).…”

Section: Methodsmentioning

confidence: 99%

Evidence That the Homeodomain Protein Gtx Is Involved in the Regulation of Oligodendrocyte Myelination

et al. 1997

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We have investigated the patterns of postnatal brain expression and DNA binding of Gtx, a homeodomain transcription factor. Gtx mRNA accumulates in parallel with the RNAs encoding the major structural proteins of myelin, myelin basic protein (MBP), and proteolipid protein (PLP) during postnatal brain development; Gtx mRNA decreases in parallel with MBP and PLP mRNAs in the brains of myelin-deficient rats, which have a point mutation in the PLP gene. Gtx mRNA is expressed in differentiated, postmitotic oligodendrocytes but is not found in oligodendrocyte precursors or astrocytes. These data thus demonstrate that Gtx is expressed uniquely in differentiated oligodendrocytes in postnatal rodent brain and that its expression is regulated in parallel with the major myelin protein mRNAs, encoding MBP and PLP, under a variety of physiologically relevant circumstances.Using a Gtx fusion protein produced in bacteria, we have confirmed that Gtx is a sequence-specific DNA-binding protein, which binds DNA sequences containing a core AT-rich homeodomain binding site. Immunoprecipitation of labeled DNA fragments encoding either the MBP or PLP promoter regions with this fusion protein has identified several Gtx-binding fragments, and we have confirmed these data using an electrophoretic mobility shift assay. In this way we have identified four Gtx binding sites within the first 750 bp of the MBP promoter and four Gtx binding sites within the first 1.3 kb of the PLP promoter. In addition, inspection of the PLP promoter sequence demonstrates the presence of six additional Gtx binding sites. These data, taken together, strongly suggest that Gtx is important for the function of differentiated oligodendrocytes and may be involved in the regulation of myelin-specific gene expression.

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“…TPO-specific probes were generated from either the 2.3-kb BamHI fragment of clone 19 (see below), the 1.6-kb EcoRI-StuI fragment of clone 18, or the PvuII-digested 5.5-kb EcoRI fragment of clone 18. In addition, templates included the 1.7-kb EcoRI fragment of pMBP1 (Saavedra et al, 1989), the 1.4-kb EcoRI fragment of pCNP4 (2 ',3 '-cyclic nucleotide 3'-phosphohydrolase; CNP) (Bernier et al, 1987), and the 0.5-kb EcoRI-Hindlil fragment of pXir40s (28S RNA) (Timsit et al, 1992).…”

Section: Rna Isolation and Analysismentioning

confidence: 99%

TPO1, a Member of a Novel Protein Family, Is Developmentally Regulated in Cultured Oligodendrocytes

Krueger

Gonye

Madison

et al. 1997

Journal of Neurochemistry

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Although the myelin membrane contains only a small set of major proteins, more sensitive assays indicate the presence of a plethora of uncharacterized proteins. We have used an antibody perturbation approach to reversibly block the differentiation of prooligodendroblasts into myelinating cells, and, in combination with a differential screening procedure, identified novel mRNAs that are activated during this period. One cDNA, TPO1, recognizes a 5.5-kb mRNA that is strongly up-regulated in oligodendrocytes after release of the differentiation block and that is expressed at high levels in brain tissue during active myelination. This cDNA represents at least two mRNAs differing from each other in their 5'-termini. The TPO1 cDNA contains an open reading frame of 1,380 bp, encoding a protein of 51 .8 kDa with a predicted pi of 9.1 that contains two regions homologous to nonclassic zinc finger motifs. Subceilular localization studies suggest the enriched presence of TPO1 in spherical structures along the major cytoplasmic processes of oligodendrocytes. TPO1, along with homologues expressed in testis, placenta, and PC12 cells, form a novel family of proteins with multiple hydrophobic domains possibly serving as membrane spanning regions. We postulate that in oligodendrocytes, TPO1 encodes a protein factor involved in myeiin biogenesis. Key Words: TPO1 -Oligodendrocyte development-Myelin-Zinc finger-Membrane protein-OB-fold.

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Molecular cloning of a 2',3'-cyclic nucleotide 3'-phosphodiesterase: mRNAs with different 5' ends encode the same set of proteins in nervous and lymphoid tissues

Cited by 137 publications

References 38 publications

Identification of Essential Residues in 2′,3′-Cyclic Nucleotide 3′-Phosphodiesterase

Identification of Essential Residues in 2′,3′-Cyclic Nucleotide 3′-Phosphodiesterase

Evidence That the Homeodomain Protein Gtx Is Involved in the Regulation of Oligodendrocyte Myelination

TPO1, a Member of a Novel Protein Family, Is Developmentally Regulated in Cultured Oligodendrocytes

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