Aging affects the distribution of the circadian CLOCK protein in rat hepatocytes

Abstract

Several biochemical, physiological, and behavioral processes exhibit cyclic oscillations of about 24 h, which have been defined as circadian rhythms. In mammals, the primary circadian pacemaker resides in the suprachiasmatic nuclei; however, cell-autonomous circadian oscillators occur also in extraneural tissues, including the liver. CLOCK protein is a transcription factor essential for normal circadian rhythms and recent studies have demonstrated that it undergoes intranuclear redistribution in hepatocytes, along the daily cycle. It is known that aging leads to a progressive deterioration of the circadian rhythm at the behavioral, physiological, and cellular levels; in addition, aging affects the organization of nuclear structural components involved in transcription and splicing. In this view, we carried out ultrastructural immunocytochemical analyses on hepatocytes of adult and old rats, so as to investigate possible qualitative and quantitative modifications of CLOCK protein, in relation to the aging process. Our observations demonstrated that most CLOCK protein was always located in the cell nucleus, where it accumulated on perichromatin fibrils (the sites of premRNA transcription and early splicing); in addition, CLOCK showed daily oscillations in the different nuclear compartments, but these oscillations differed significantly between adult and old animals. This unusual distribution of CLOCK protein during aging could be related to the prolonged diurnal activity of old animals and/or to altered nuclear pathways.

Influence of Temperature on the Liver Circadian Clock in the Ruin Lizard Podarcis sicula

“…Finally, the anti-CLOCK signal occurring in the cytoplasm, in particular on the rough endoplasmic reticulum, probably revealed synthesized molecules before entering the nucleus. Interestingly, the intracellular distribution of lCLOCK protein observed in lizards is quite similar to that previously observed in different mammalian species (Malatesta et al, 2003a(Malatesta et al, ,b, 2005. Moreover, the hepatocyte nuclei of hypothermic lizards contain nuclear bodies reminding the HERDS, structural constituents forming by RNP segregation when transcription decreases (Biggiogera and Pellicciari, 2000;Biggiogera et al, 2004).…”

Effects of ageing on the fine distribution of the circadian CLOCK protein in reticular formation neurons

“…The ultrastructural distribution of CLOCK protein observed in the neurons of the reticular formation is comparable to that previously observed in hepatocytes of rats (Malatesta et al 2005) as well as of other species (Malatesta et al 2003a(Malatesta et al , b, 2007. CLOCK is a bHLH-PAS transcription factor (Reppert and Weaver 2001) and, therefore, preferentially associates with both PF (i.e., the in situ form of pre-mRNA transcription and early splicing: Fakan 1994Fakan , 2004, and the nucleolar dense Wbrillar component (i.e., the site of rRNA transcription and early splicing: Cmarko et al 2000;Biggiogera et al 2001, Fakan 2004).…”

“…The nucleolus showed some labelling on its dense Wbrillar component. In both adult and old rats no modiWcation of any cytoplasmic or nuclear structural constituent was observed in the diVerent daily phases considered, according to previous observations in rat, mouse and lizard (Malatesta et al 2003a(Malatesta et al , 2005(Malatesta et al , 2007.…”

“…Similarly to rat hepatocytes (Malatesta et al 2005), CLOCK protein in the reticular formation neurons of adult animals maintains the same intracellular distribution pattern during the daily cycle and shows quantitative daily oscillations, with a peak at ZT12, in all cellular compartments considered (apart from IG). In rat hepatocytes the peak occurs at ZT4 (Malatesta et al 2005), but this shift could be related to tissue-speciWc diVerences (Morse and SassoneCorsi 2002;Gachon et al 2004).…”

“…Moreover, it has been shown that age alters the daily expression of CLOCK and BMAL1 proteins in the hamster suprachiasmatic nuclei (Kolker et al 2003) as well as of per1 and per2 genes in aged rats and mice (Weinert et al 2001;Yamazaki et al 2002). Finally, alterations of the intracellular distribution of CLOCK protein have been described in peripheral tissues of old rats (Malatesta et al 2005).…”