Molecular mechanisms of proteasome plasticity in aging

Abstract

The ubiquitin-proteasome pathway plays a crucial role in regulation of intracellular protein turnover. Proteasome, the central protease of the pathway, encompasses multisubunit assemblies sharing a common catalytic core supplemented by regulatory modules and localizing to different subcellular compartments. To better comprehend age-related functions of the proteasome we surveyed content, composition and catalytic properties of the enzyme in cytosolic, microsomal and nuclear fractions. obtained from mouse livers subjected to organismal aging. We found that during aging subunit composition and subcellular distribution of proteasomes changed without substantial alterations in the total level of core complexes. We observed that the general decline in proteasomes functions was limited to nuclear and cytosolic compartments. Surprisingly, the observed changes in activity and specificity were linked to the amount of the activator module and distinct composition of the catalytic subunits. In contrast, activity, specificity and composition of the microsomal-associated proteasomes remained mostly unaffected by aging; however their relative contribution to the total activity was substantially elevated. Unexpectedly, the nuclear proteasomes were affected most profoundly by aging possibly triggering significant changes in cellular signaling and transcription. Collectively, the data indicate an age-related refocusing of proteasome from the compartment specific functions towards general protein maintenance.

Molecular alterations in proteasomes of rat liver during aging result in altered proteolytic activities

“…Magnification, ×40 different proteases (Puente and Lopez-Otin 2004), which could be differently affected by aging or protein extraction protocols and therefore mask the real age-dependent alteration of proteasome activities. Indeed, our analysis of 20S proteasomes purified to apparent homogeneity and measurement of their peptide hydrolysing activity using fluorogenic peptide substrates revealed a significantly decreased caspase-like activity in 20S proteasome population purified from livers of aged as compared to young rats, which is in agreement with results of other investigators (Shibatani et al 1996;Conconi et al 1996;Hayashi and Goto 1998;Petersen et al 2010;Rodriguez et al 2010). This phenomenon most likely not only reflects alterations in the specific activities of the 20S proteasome subpopulations but also in their concentrations, keeping in mind that each subpopulation has a specific pattern of activities.…”

“…Such a change in proteasome content did not correspond to a decrease of the chymotryptic, tryptic and caspase-like activities in total liver extracts of aged rats. To evaluate this in correlation to contrasting results of other studies (Breusing et al 2009;Dasuri et al 2009;Hayashi and Goto 1998;Rodriguez et al 2010), we have to consider that hydrolysis of short fluorogenic peptides in crude tissue extracts always reflects the sum of activities of Fig. 7 Immunohistochemical localisation of proteasome-subunits in rat liver.…”

“…Their activity towards short fluorogenic peptide substrates is different, an aspect taken into consideration in some of the investigations. In some investigations, a parallel decrease of 20S and 26S proteasomes was found in liver of aging rat (Hayashi and Goto 1998;Dasuri et al 2009), whereas a decline of PA28 and 19S regulator (Rodriguez et al 2010) or just of the 19S regulator subunit Rpn10 was suggested to be responsible for a decreasing activity during aging (Huber et al 2009). …”

“…Their stoichiometry is α 7 β 7 β 7 α 7 , and three of the β-subunits, β1, β2 and β5, contain the proteolytically active sites catalysing the caspase-, trypsin-, and chymotrypsin-like activities, respectively. In liver tissue of aging mice and rats, all or some of these activities when measured with fluorogenic peptide substrates have been found to decrease (Sahakian et al 1995;Shibatani et al 1996;Conconi et al 1996;Breusing et al 2009;Dasuri et al 2009;Hayashi and Goto 1998;Rodriguez et al 2010) or decrease temporarily before increasing again (Petersen et al 2010;Keller et al 2000). However, no change of activity was measured in liver of aged as compared to young mice in one investigation (Huber et al 2009), an observation corroborating the data published by Scrofano and colleagues, who determined liver proteasome activity in young and old mice by use of β-lactoglobulin and oxidized ribonuclease as substrates (Scrofano et al 1998).…”

“…Activity measurements of proteasomes in all of these investigations were performed in tissue extracts or with extracts partially fractionated by chromatographic or centrifugation steps (Hayashi and Goto 1998;Rodriguez et al 2010). Only Conconi et al (1996) measured the activity of 20S proteasomes after their purification to apparent homogeneity.…”