Abstract
IntroductionNuclear accumulation of a mutant form of the nuclear protein Lamin-A, called Progerin (PG) or Lamin AΔ50, occurs in Hutchinson-Gilford Progeria Syndrome (HGPS) or Progeria, an accelerated aging disease. One of the main symptoms of this genetic disorder is a loss of sub-cutaneous fat due to a dramatic lipodystrophy.MethodsWe stably induced the expression of human PG and GFP -Green Fluorescent Protein- as control in 3T3L1 cells using a lentiviral system to study the effect of PG expression in the differentiation capacity of this cell line, one of the most used adipogenic models. Quantitative proteomics (iTRAQ) was done to study the effect of the PG accumulation. Several of the modulated proteins were validated by immunoblotting and real-time PCR. Mitochondrial function was analyzed by measurement of a) the mitochondrial basal activity, b) the superoxide anion production and c) the individual efficiency of the different complex of the respiratory chain.ResultsWe found that over-expression PG by lentiviral gene delivery leads to a decrease in the proliferation rate and to defects in adipogenic capacity when compared to the control. Quantitative proteomics analysis showed 181 proteins significantly (p < 0.05) modulated in PG-expressing preadipocytes. Mitochondrial function is impaired in PG-expressing cells. Specifically, we have detected an increase in the activity of the complex I and an overproduction of Superoxide anion. Incubation with Reactive Oxygen Species (ROS) scavenger agents drives to a decrease in autophagic proteolysis as revealed by LC3-II/LC3-I ratio.ConclusionPG expression in 3T3L1 cells promotes changes in several Biological Processes, including structure of cytoskeleton, lipid metabolism, calcium regulation, translation, protein folding and energy generation by the mitochondria. Our data strengthen the contribution of ROS accumulation to the premature aging phenotype and establish a link between mitochondrial dysfunction and loss of proteostasis in HGPS.
Highlights
Nuclear accumulation of a mutant form of the nuclear protein Lamin-A, called Progerin (PG) or Lamin AΔ50, occurs in Hutchinson-Gilford Progeria Syndrome (HGPS) or Progeria, an accelerated aging disease
Peinado et al [40] demonstrated using a differential in gel electrophoresis (DIGE) approach that mitochondrial dysfunction and disorganization of the cytoskeleton were characteristic of the adipose tissue in ZMPSTE24 null mice, which mimic some but not all of the symptoms associated with HGPS-derived cells
We have found a dramatic increase of this group of proteins in PG-3T3L1 cells, indicating an activation of chaperone-mediated autophagy (CMA), a process recently described in age-related pathologies [58, 59]
Summary
Nuclear accumulation of a mutant form of the nuclear protein Lamin-A, called Progerin (PG) or Lamin AΔ50, occurs in Hutchinson-Gilford Progeria Syndrome (HGPS) or Progeria, an accelerated aging disease. Mutations in the LMNA gene are the causal agent for a subset of genetic diseases affecting mesoderm tissues called laminopathies [1]. Historically thought to be involved only in nuclear structure, roles in replication, chromatin organization and stem cell differentiation have been demonstrated recently for Lamin A [8, 9]. Farnesylated PG does not properly process, accumulates in the nuclear envelope, causes structural defects in the nuclear lamina and may be interfering with regulation of the signalling pathway mediated by p16/Rb necessary to maintain the balance between differentiation and proliferation of stem cells in the tissue regeneration process [8]. Several studies showed the importance of accumulation of the farnesylated precursor in the development of the disease [11,12,13]
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