Circadian clocks regulate adenylyl cyclase activity rhythms in human RPE cells.
Genes and components of the circadian clock may represent relevant drug targets for diseases involving circadian dysfunctions. By exploiting an established cell line derived from human retinal pigment epithelium (HRPE), the cell constituting the blood-retinal barrier that is essential to maintain the visual functions of the sensorineural retina, we showed serum-shock induction of rhythmic changes in forskolin-evoked adenylyl cyclase (AC) activity. In the presence of Ca2+ and protein kinase A, the forskolin-induced AC activity is significantly, but not completely inhibited, suggesting the involvement of both Ca2+-sensitive and Ca2+-insensitive AC isoforms in the regulation of circadian rhythmicity in these cells. Semi-quantitative RT-PCR showed circadian profile in the expression of three AC isoforms, the Ca2+-inhibitable AC5 and AC6 and the Ca2+-insensitive AC7, and the clock genes hPer1 and hPer2. Our results demonstrate for the first time circadian rhythmicity in a human cell line, identifying the isoforms involved in the circadian profile of AC activity and showing a rhythmicity of the clock gene mRNA expression in these cells. Therefore, the results reported here provide evidence for an intertwine between AC/[Ca2+]i signalling pathways and Per genes in the HRPE circadian clockwork.
Pavan
Barbara
B
Department of Biology, Section of General Physiology, University of Ferrara, via L. Borsari, 46, 44100-I Ferrara, Italy.
Frigato
Elena
E
Pozzati
Sara
S
Prasad
Puttur D
PD
Bertolucci
Cristiano
C
Biondi
Carla
C
D000262:Adenylyl Cyclases / Q000235:genetics* / Q000378:metabolism*; D001683:Biological Clocks / Q000502:physiology*; D018797:Cell Cycle Proteins / Q000235:genetics; D002462:Cell Membrane / Q000201:enzymology; D002478:Cells, Cultured; D005786:Gene Expression Regulation; D006801:Humans; D009687:Nuclear Proteins / Q000235:genetics; D056950:Period Circadian Proteins; D010857:Pigment Epithelium of Eye / Q000201:enzymology*; D014157:Transcription Factors / Q000235:genetics
D018797:Cell Cycle Proteins; D009687:Nuclear Proteins; C539022:PER1 protein, human; C421902:PER2 protein, human; D056950:Period Circadian Proteins; D014157:Transcription Factors; D000262:Adenylyl Cyclases