Specific use of start codons and cellular localization of splice variants of human phosphodiesterase 9A gene.
BACKGROUND
Phosphodiesterases are an important protein family that catalyse the hydrolysis of cyclic nucleotide monophosphates (cAMP and cGMP), second intracellular messengers responsible for transducing a variety of extra-cellular signals. A number of different splice variants have been observed for the human phosphodiesterase 9A gene, a cGMP-specific high-affinity PDE. These mRNAs differ in the use of specific combinations of exons located at the 5' end of the gene while the 3' half, that codes for the catalytic domain of the protein, always has the same combination of exons. It was observed that to deduce the protein sequence with the catalytic domain from all the variants, at least two ATG start codons have to be used. Alternatively some variants code for shorter non-functional polypeptides.


RESULTS
In the present study, we expressed different splice variants of PDE9A in HeLa and Cos-1 cells with EGFP fluorescent protein in phase with the catalytic domain sequence in order to test the different start codon usage in each splice variant. It was found that at least two ATG start codons may be used and that the open reading frame that includes the catalytic domain may be translated. In addition the proteins produced from some of the splice variants are targeted to membrane ruffles and cellular vesicles while other variants appear to be cytoplasmic. A hypothesis about the functional meaning of these results is discussed.


CONCLUSIONS
Our data suggest the utilization of two different start codons to produce a variety of different PDE9A proteins, allowing specific subcellular location of PDE9A splice variants.
Rentero
Carles
C
Departament de Genètica Molecular, Institut de Biologia Molecular de Barcelona, CSIC, Jordi Girona, 18, 08034 Barcelona, Spain. c.rentero@unsw.edu.au <c.rentero@unsw.edu.au>
Puigdomènech
Pere
P
D015105:3',5'-Cyclic-AMP Phosphodiesterases / Q000096:biosynthesis / Q000737:chemistry / Q000235:genetics / Q000502:physiology*; D000818:Animals; D015153:Blotting, Western / Q000379:methods; D019556:COS Cells; D002462:Cell Membrane / Q000201:enzymology; D002522:Chlorocebus aethiops; D018387:Codon, Initiator / Q000235:genetics*; D015971:Gene Expression Regulation, Enzymologic / Q000235:genetics*; D023061:Gene Order / Q000235:genetics; D006056:Golgi Apparatus / Q000201:enzymology; D006367:HeLa Cells; D006801:Humans; D007527:Isoenzymes / Q000096:biosynthesis / Q000737:chemistry / Q000235:genetics / Q000502:physiology; D018613:Microscopy, Confocal / Q000379:methods; D008969:Molecular Sequence Data; D015398:Signal Transduction / Q000235:genetics; D014162:Transfection
D018387:Codon, Initiator; D007527:Isoenzymes; D015105:3',5'-Cyclic-AMP Phosphodiesterases; C116137:PDE9A protein, human