Regulation of autophagy in mammalian cells by stress activated signaling pathways
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Date
2013-04-02
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Abstract
Autophagy is an intracellular bulk degradation process of cytoplasmic proteins and
organelles aimed to preserve cellular homeostasis. It involves the formation of
double-membrane structures, called autophagosomes that transport their cargo to
the lysosomes where it becomes degraded. The molecular mechanisms underlying
induction of autophagy, biogenesis, trafficking and turnover of autophagosomes
have been extensively studied. However, the regulation of these steps by signaling
pathways that adjust the autophagy flux to different cellular stress situations is still
poorly understood. The aim of this work was to investigate the regulation of
autophagy upon particular stress conditions in a time-dependent manner. We
particularly focused on the control of autophagy by p38. For this purpose we applied
a MCF-7/NeuT cell model of oncogene-induced senescence (OIS). In this model,
doxycycline-inducible expression of an oncogenic variant of ERBB2 (NeuT) in the
breast cancer cell line MCF-7 leads to premature senescence, a process that was
shown to be accompanied by aberrant accumulation of autophagic vesicles. We
found that autophagy becomes initially induced but that its maturation is later
impaired upon oncogenic stress. In addition we identified p38/MAPK as the signaling
pathway mediating the block of autophagic flux. This was evidenced by the fact that
pharmacological inhibition of p38 by the small molecule compound SB203580 can
restore the autophagic flux. This result was confirmed in time-dependent rescue
experiments with SB203580. Furthermore, mechanisms by which p38 mediates the
autophagic flux blockade were identified. These involved post-translational
modifications of the microtubule-associated proteins MAP4 and OP18/STMN1, as
well as down regulation of a particular dynein motor protein subunit DYNLL1, which
all together provoke microtubule disruption and defective autophagosome transport
to the lysosome. The biphasic behaviour of autophagy upon stress (early induction,
late block) was confirmed in several cell lines exposed to a different stress-stimulus
(anisomycin). Here we could show that also p38/MAPK is involved in these effects in
a dual way, with an early induction of initial steps of autophagy and a later block of
maturation. In addition, the p38-mediated block of the autophagic maturation was
shown to occur via the same mechanism described above.In conclusion, this work revealed new aspects of the control of autophagy by p38
signaling, mainly that the duration and intensity of p38 activation are key
determinants of the autophagic flux rate. Low or transitory stress conditions that
moderately activate p38 stimulate the autophagic flux. In contrast, more permanent
and intensive stress that causes a strong activation of p38 blocks the autophagic
flux.
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Keywords
Autophagy, Microtubules, p38, Senescence