CNP2 mRNA directs synthesis of both CNP1 and CNP2 polypeptides

Abstract

The ribosome scanning model for translational initiation predicts that eukaryotic mRNAs should, as a rule, be monocistronic. However, cases have recently been described of eukaryotic mRNAs producing more than one protein through alternative translational initiation at several different AUG codons. The present work reports the occurrence of two translational start sites on the mRNA encoding isoform 2 of the myelin marker enzyme 2',3'-cyclic nucleotide 3'-phosphodiesterase (CNP) in rat and mouse. We show that the CNP2 mRNA is able to direct synthesis of not only CNP2, but also CNP1 polypeptide. Immunoprecipitation experiments using a polyclonal antibody directed against CNP detect both CNP isoforms in tissues or cell lines expressing only the CNP2 transcript. Thus, the synthesis of CNP1 and CNP2 polypeptides must be encoded by the CNP2 transcript. In vitro translation of synthetic CNP2 mRNA demonstrates that both CNP isoforms are synthesized by initiation at different AUG codons. Furthermore, by introducing mutations to "switch off" translation from the second in-frame AUG codon in the CNP2 cDNA, and transfecting 293T cells with those constructs, we are able to correlate the production of CNP1 and CNP2 with different translational start sites. These results lead us to conclude that the CNP2 mRNA is able to produce both CNP1 and CNP2 polypeptides. This investigation has altered our understanding of the temporal expression of the CNP protein isoforms during development of the central nervous system (CNS).

Oligodendrocyte progenitor cells can act as cell of origin for experimental glioma

“…Several studies have shown a specific expression of CNP during differentiation of oligodendrocytes (Scherer et al, 1994;Yuan et al, 2002) and it has been proven to faithfully direct expression of a transgene to OPCs and differentiated oligodendrocytes (Gravel et al, 1998;Chandross et al, 1999;Yuan et al, 2002). The Cnp promoter consists of two separate promoter regions resulting in two isoforms, Cnp1 (46 kDa) and Cnp2 (48 kDa), encoded by the same gene (O'Neill et al, 1997). The proximal promoter, which regulates transcription of Cnp1 mRNA, contains one open-reading frame for CNP1 protein and is mainly active after birth.…”

“…The proximal promoter, which regulates transcription of Cnp1 mRNA, contains one open-reading frame for CNP1 protein and is mainly active after birth. The distal promoter regulates transcription of Cnp2 mRNA, which produces both CNP1 and CNP2 polypeptides and is active both in embryonic brain and postnatally (O'Neill et al, 1997). The transgenic construct contained the entire mouse Cnp promoter region coupled to the Tv-a gene followed by an ires (internal ribosomal entry site) sequence and the lacZ gene ( Figure 1a).…”

Ca2+-dependent permeability transition regulation in rat brain mitochondria by 2′,3′-cyclic nucleotides and 2′,3′-cyclic nucleotide 3′-phosphodiesterase

“…Two CNP isoforms, CNP1 (46 kDa) and CNP2 (48 kDa), were found (16,23) that are encoded by a single gene. The two isoforms are due to the presence of two alternative translation start sites (28). Several posttranslational modifications of CNP, such as phosphorylation, isoprenylation, and acylation, with unclear functional consequences are known (1,2,10,36,38,39).…”

Nonspecific association of 2',3'-cyclic nucleotide 3'-phosphodiesterase with the rat forebrain postsynaptic density fraction

“…The CNP is a protein of unknown function that is abundantly expressed in myelinating cells (Scherer et al, 1994;O'Neill et al, 1997). The CNP is also expressed at low level in both neuronal and nonneuronal tissues as described in Introduction.…”

“…The 2',3'-cyclic nucleotide 3'-phosphodiesterase (CNP; EC 3.1.4.37) is expressed as two isoforms with an apparent molecular mass of 46 kDa (CNP1) and 48 kDa (CNP2), which are produced by alternative ribosomal initiation at two different AUG codons, thus differing each other only by the 20-amino acid extension at the N terminus (O'Neill et al, 1997). Despite its lengthy history, relatively little is known about the function of CNP.…”