Genetic analysis of four novel peroxisome proliferator activated receptor-gamma splice variants in monkey macrophages.
Peroxisome proliferator activated receptor-gamma (PPAR-gamma) is abundantly expressed in atherosclerotic lesions and is implicated in atherogenesis. The existence of three splice variants, PPAR-gamma 1, PPAR-gamma 2, and PPAR-gamma 3 has been established. Using monocyte-derived macrophages from cynomolgus monkeys, we demonstrate here the identification of two new PPAR-gamma exons, exon C and exon D, which splice together with already established exons A1, A2, and B in the 5(') terminal region to generate four novel PPAR-gamma subtypes, PPAR-gamma 4, -gamma 5, -gamma 6, and -gamma 7. PPAR-gamma 4 and gamma 5 were detected only in macrophages whereas gamma 6 and gamma 7 were expressed both in macrophages and adipose tissues. None of these novel isoforms were detected in muscle, kidney, and spleen from monkeys. We found sequences identical to exons C and D in the human genome database. These and all PPAR-gamma exons known to date are encoded by a single gene, located from region 10498 K to 10384 K on human chromosome 3. We cloned and expressed PPAR-gamma 1, PPAR-gamma 4, and PPAR-gamma 5 proteins in yeast using the expression vector pPICZB. As expected, all recombinant proteins showed a molecular weight of approximately 50 kDa. We also investigated the effect of a high-fat diet on the level of macrophage PPAR-gamma expression in monkeys. RT-PCR showed a significant increase in total PPAR-gamma and ABCA1 mRNA levels in macrophages of fat-fed monkeys (n=7) compared to those maintained on a normal diet (n=2). However, none of the novel isoforms seemed to be induced by fat-feeding. We used tetracycline-responsive expression vectors to obtain moderate expression of PPAR-gamma 4 and -gamma 5 in CHO cells. In these cells, expression of PPAR-gamma 5 but not -gamma 4 repressed the expression of ABCA1. Neither isoform modulated the expression of lipoprotein lipase. Our results suggest that individual PPAR-gamma isoforms may be responsible for unique tissue-specific biological effects and that PPAR-gamma 4 and -gamma 5 may modulate macrophage function and atherogenesis.
Zhou
Jiming
J
Department of Internal Medicine, University of Iowa College of Medicine, Iowa City, IA 52242, USA.
Wilson
Katina M
KM
Medh
Jheem D
JD
D017398:Alternative Splicing; D000818:Animals; D017931:DNA Primers; D005091:Exons; D014644:Genetic Variation; D000882:Haplorhini; D006801:Humans; D008252:Macaca fascicularis; D008253:Macaca mulatta; D051379:Mice; D020033:Protein Isoforms / Q000235:genetics; D018160:Receptors, Cytoplasmic and Nuclear / Q000235:genetics*; D013552:Swine; D014157:Transcription Factors / Q000235:genetics*
D017931:DNA Primers; D020033:Protein Isoforms; D018160:Receptors, Cytoplasmic and Nuclear; D014157:Transcription Factors