Ž . Biochimica et Biophysica Acta 1353 1997 287–297 Multiple splice variants of phosphodiesterase PDE4C cloned from human lung and testis Rena Obernolte, James Ratzliff, Preston A. Baecker, Donald V. Daniels, Patti Zuppan 1 , Kurt Jarnagin ) , Earl R. Shelton 2 Roche Bioscience, 3401 HillÕiew AÕe., S3-1, Palo Alto, CA 94304, USA Received 3 April 1997; accepted 28 April 1997 Abstract Ž . Four closely related cyclic-nucleotide specific phosphodiesterase PDE4 genes have been identified in both humans and rats: PDE4A, 4B, 4C and 4D. We have now cloned cDNAs for multiple splice variants of human PDE4C. Two splice Ž . variants, PDE4C-791 and PDE4C-426, were isolated from a fetal lung library. The longest open reading frame ORF of Ž . w 791 amino acids aa is encoded by PDE4C-791, which is similar to a recently described cDNA Engels, P., Sullivan, M., Ž . x X Muller, T. and Lubbert, H. FEBS Lett. 358 1995 305–10 , except that an alternative 5 -end sequence upstream of the first ¨ methionine extends the PDE4C-791 ORF by 79 aa. The PDE4C-426 variant contains 3 insertions that are located 5 X to the catalytic domain and encode several in-frame stop codons. The predicted 426 aa protein initiates at a methionine 365 aa within PDE4C-791. A baculovirus clone starting at this methionine expressed an enzymatically active protein. Two additional splice variants, PDE4C-D54 and PDE4C-D109, were found in testis mRNA. PDE4C-D54 contained a novel 5 X -end region and a deletion of 162 nt; the predicted protein deletes 54 aa from the amino-terminal region. The PDE4C-D54 protein produced in baculovirus-infected cells was enzymatically active and sensitive to PDE4-specific inhibitors. The PDE4C-D109 protein is similar to PDE4C-D54 but has an additional 55 aa deleted in the catalytic domain; it lacked Ž . enzymatic activity. Analysis of uncloned total mRNA from 4 tissue sources by polymerase chain reaction PCR confirmed the presence of mRNAs with the two deletions and three insertions that we observed in cDNA clones. The PDE4C-D54 Ž. Ž. Abbreviations: aa, amino acid s ; Ap, ampicillin; bp, base pair s ; cDNA, DNA complementary to RNA; cfu, colony forming unit; Ž. Ž. Ž. EtdBr, ethidium bromide; kb, kilobase s or 1000 bp; kDa, kilodalton s ; nt, nucleotide s ; ORF, open reading frame; PCR, polymerase chain reaction; PDE, cyclic-nucleotide phosphodiesterase – nomenclature for PDE genes is taken from the most recent recommendations wx Ž . 4 , and in the case of cDNAs a number referring to the total aa length of the predicted ORF PDE4C-791, PDE4C-426 or to the total Ž . size of aa deletions within the ORF PDE4C-D54, and PDE4C-D109 is appended; PDE4C, phosphodiesterase 4C protein; PDE4C, Ž . gene encoding PDE4C; PDE4C-791, nucleic acid RNA, DNA from an alternatively spliced PDE4C mRNA; pfu, plaque-forming unit; w x SM, suspension medium 17 ; SSC, 0.15 M NaClr0.015 M Na P citrate pH 7.6 3 ) Ž . Corresponding author. Fax: q1 415 3547554. E-mail: kurt.jarnagin@roche.com 1 Present address: Biotech Consultant, San Jose, CA 95125, USA. 2 Present address: Kowa Research Institute, Inc., 1731 Technology Dr. a460, San Jose, CA 95110, USA. E-mail: eshelton@best.com 0167-4781r97r$17.00 q 1997 Elsevier Science B.V. All rights reserved. Ž . PII S0167-4781 97 00080-8