Abstract
The aging process is a physiological phenomenon associated with progressive changes in metabolism, genes expression, and cellular resistance to stress. In neurons, one of the hallmarks of senescence is a disturbance of calcium homeostasis that may have far-reaching detrimental consequences on neuronal physiology and function. Among several proteins involved in calcium handling, plasma membrane Ca2+-ATPase (PMCA) is the most sensitive calcium detector controlling calcium homeostasis. PMCA exists in four main isoforms and PMCA2 and PMCA3 are highly expressed in the brain. The overall effects of impaired calcium extrusion due to age-dependent decline of PMCA function seem to accumulate with age, increasing the susceptibility to neurotoxic insults. To analyze the PMCA role in neuronal cells, we have developed stable transfected differentiated PC12 lines with down-regulated PMCA2 or PMCA3 isoforms to mimic age-related changes. The resting Ca2+ increased in both PMCA-deficient lines affecting the expression of several Ca2+-associated proteins, i.e., sarco/endoplasmic Ca2+-ATPase (SERCA), calmodulin, calcineurin, GAP43, CCR5, IP3Rs, and certain types of voltage-gated Ca2+ channels (VGCCs). Functional studies also demonstrated profound changes in intracellular pH regulation and mitochondrial metabolism. Moreover, modification of PMCAs membrane composition triggered some adaptive processes to counterbalance calcium overload, but the reduction of PMCA2 appeared to be more detrimental to the cells than PMCA3.
Highlights
The aging process is a physiological phenomenon affecting all living organisms and is associated with perpetual changes in metabolic control, the expression and stability of genes, and cellular resistance to stress
Taking into account higher pHmito in plasma membrane Ca2+-ATPase (PMCA)-deficient cells, increased accumulation of Ca2+ by mitochondria consisted with the observation of over-activation of electron transport chain (ETC) complexes, our results point to a specific coupling between PMCA2 generated calcium fluxes and metabolic activity and may mirror specific cellular alterations observed in early aging
The second important player and limiting factor was the amount of CaM available for binding. This could profoundly interfere with CaN/NFAT activation in both PC12-deficient lines, further suggesting the existence of the feedback mechanism by which CaM could affect its own expression. This specific regulation seems to be a direct consequence of selective PMCA isoform silencing, because no similar effect was observed in the control PC12 cells
Summary
The aging process is a physiological phenomenon affecting all living organisms and is associated with perpetual changes in metabolic control, the expression and stability of genes, and cellular resistance to stress. It is well established that deficits in ATP supply, which is critical for reestablishment of ion balance by the plasma membrane Ca2+-ATPase, can produce diverse defects in brain function This posits disturbed calcium homeostasis in the center of several age-associated brain pathologies and should be considered as an important risk factor for the development of neurodegenerative diseases. PMCA, by interacting locally with a number of protein partners, can be incorporated into intracellular signaling pathways [27] It suggests that PMCA may play a role that is far beyond its classically attributed function of a calcium transporter. Whereas in undifferentiated PC12 cells, all main PMCA isoforms have been detected, with PMCA4b constituting a major one, the differentiation process stimulates the expression of additional splice variants (i.e. 1c, 2a, 2c, 4a) with unknown physiological function [47,48]. Our extensive functional and molecular analyses gave new insights into downstream cellular processes that can be modified by altered PMCA composition
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