Protein damage segregates asymmetrically in dividing yeast cells, rejuvenating daughters at the expense of mother cells. Zhou et al. now show that newly synthesized proteins are particularly prone to aggregation and describe a mechanism that tethers aggregated proteins to mitochondria. This association constrains aggregate mobility, effectively retaining and sorting toxic aggregates away from younger cells

Bukau, Bernd

Mogk, Axel

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PreviewsMitochondria Tether Protein Trash
to Rejuvenate Cellular Environments
Axel Mogk1 and Bernd Bukau1,*
1Zentrum fu¨r Molekulare Biologie der Universita¨t Heidelberg (ZMBH) and Deutsches Krebsforschungszentrum (DKFZ), DKFZ-ZMBHAlliance,
Im Neuenheimer Feld 282, 69120 Heidelberg, Germany
*Correspondence: bukau@zmbh.uni-heidelberg.de
http://dx.doi.org/10.1016/j.cell.2014.10.007
Protein damage segregates asymmetrically in dividing yeast cells, rejuvenating daughters at the
expense of mother cells. Zhou et al. now show that newly synthesized proteins are particularly
prone to aggregation and describe a mechanism that tethers aggregated proteins to mitochondria.
This association constrains aggregate mobility, effectively retaining and sorting toxic aggregates
away from younger cells.Accumulation of damaged macromole-
cules accompanies aging in all organisms
and is considered a major factor contrib-
uting to degeneration of cells and tis-
sues. Asymmetric distribution of molecu-
lar debris during cell division represents
a conserved cellular strategy limiting the
spread of protein damage over time and
minimizing levels of damage inherited
by younger cells. This process is best
studied in the baker’s yeast Saccharo-
myces cerevisiae, in which asymmetric
segregation during bud formation is docu-
mented for extrachromosomal rDNA
circles (ERCs), oxidatively damaged (car-
bonylated) proteins, and protein aggre-
gates composed of misfolded con-
formers. These species accumulate in
aged cells or during proteotoxic stress.
Pioneering work by the Nystrom group
shows that carbonylated proteins and
protein aggregates are largely retained
in mother cells, freeing younger cells
(daughter buds) of damage and resetting
replicative potential in the bud (Aguilaniu
et al., 2003; Liu et al., 2010). Disruption
of asymmetric damage inheritance in
yeast mutants lacking either the Sirtuin
Sir2, a factor centrally involved in aging,
or the disaggregase Hsp104, which effi-
ciently resolves aggregates after stress,
correlates with shorter replicative life-
span, reflected by fewer cell divisions
prior to senescence (Erjavec et al.,
2007). Asymmetric segregation of cellular
trash is therefore linked, though indirectly,
to daughter rejuvenation at the expense of
the older mother cell. In this issue, Zhou
et al. (2014) describe a direct link betweenprotein trash sorting and mitochondrial
inheritance.
The mechanism of damage retention
has been the focus of intense research
and controversy, resulting in conflicting
models. In one model, the retention of
carbonylated (Erjavec et al., 2007; Tes-
sarz et al., 2009) and aggregated (Liu
et al., 2010) proteins relies on a functional
actin cytoskeleton. Nystrom and co-
workers show that protein aggregates
are tethered to the actin cytoskeleton.
Retrograde flow of actin cables, nucle-
ating at the polarisome at the tip of
daughter cells, either prevents aggre-
gates entering the bud or clears the bud
from aggregates. This is an active process
involving the actin-associated motor pro-
tein Myo2 and Hsp104 (Liu et al., 2010;
Song et al., 2014) (Figure 1). Li and co-
workers, however, used quantitative par-
ticle tracking in yeast cells and failed to
show any directional bias in aggregate
movement (Zhou et al., 2011). Instead,
they observed a random walk of aggre-
gates with some degree of confinement.
The inference is that the constrained
mobility of aggregates and narrowness
of the bud neck combined are sufficient
to retain aggregated proteins in the
mother cell, suggesting an essentially
passive process. The work published by
Zhou et al. now broaches two related
key issues: first, the source of aggre-
gating proteins and second, the basis of
confined aggregate mobility.
This work pinpoints newly synthesized
proteins that, upon proteotoxic stress,
become tethered to mitochondria (andCell 159initially also the endoplasmic reticulum)
for retention in the mother cell. The
experimental model used involves pro-
tein aggregation triggered by heat shock
and visualized by the GFP-tagged dis-
aggregase Hsp104, which allows the
tracking of endogenous aggregates while
dispensing with specific misfolded pro-
tein reporters. Remarkably, inhibition of
protein synthesis abrogates formation
of detectable aggregates, even in the
case of thermolabile proteins that mis-
fold at increased temperatures. This indi-
cates that newly synthesized proteins
are particularly vulnerable to proteotoxic
stress, represent a major source of aggre-
gating protein species, and determine
the site of initial aggregation of unstable
proteins.
With regard to aggregate mobility,
fluorescence microscopy and serial
sectioning electron microscopy analyses
carried out by Zhou et al. reveal that
protein aggregates specifically associate
with endoplasmic reticulum (ER) and
mitochondria. Most protein aggregates
(90%) appear to form initially at the sur-
face of the ER. Here, aggregates also
become linked to mitochondria and
frequently appear first at ER-mitochon-
dria contact sites distinct from ERMES
(Figure 1). Aggregate capture by mito-
chondria increases upon stress relief,
suggesting a sustained aggregate-trap-
ping function of mitochondria.
Zhou et al. show that the tethering
of protein aggregates constrains aggre-
gate mobility, which also prevents
aggregate segregation to daughter cells., October 23, 2014 ª2014 Elsevier Inc. 471
Figure 1. Asymmetric Inheritance of Aggregated Proteins in Yeast
(A and B) Protein aggregates are either tethered to actin cables (A) or endoplasmic reticulum (ER) and
mitochondria organelles (B). Retrograde flow of actin cables retains aggregated proteins in the mother cell
or clears the bud from aggregates (A). The association of aggregates with mitochondria constrains
aggregate mobility and prevents their leakage to daughter cells (B).Mitochondria, however, are dynamic
organelles that rapidly extend into the
outgrowing bud during cytokinesis and
cell division. This at first glance presents
a paradox, but the authors show that
bud-inherited mitochondria are largely
devoid of aggregates. Older and more
oxidized mitochondria are also preferen-
tially retained (Higuchi et al., 2013), sug-
gesting a convergence of functions. This
supports the broader concept of an inter-
related cellular program for segregating
damage away from younger cells.
A limited genetic screen provides
further evidence that tethering to mito-
chondria contributes to asymmetric in-
heritance of aggregates. Decreased as-
sociation of mitochondria with protein
aggregates is linked to higher aggregate
mobility and reduced retention in the
mother. This profile is displayed by yeast
cells lacking Fis1, a factor required for
mitochondrial fission. Aggregates do not
colocalize with sites of mitochondrial
fission, however, hinting that Fis1 might
have a further, previously unrecognized
function relating to aggregate tethering.
However, several different yeast fission
mutants, including fis1D cells, exhibit a
slightly increased replicative lifespan
(Scheckhuber et al., 2007) despite segre-
gating protein aggregates more evenly.
The relationship between protein dam-
age segregation and lifespan is therefore
more complex than suggested by current
models.
Finally, naturally formed or heat-stress-
induced aggregates no longer colocalize
with mitochondria in aged yeast cells,472 Cell 159, October 23, 2014 ª2014 Elsevisuggesting that aggregate sorting deteri-
orates with age. This seems a chicken-
and-egg question, as it is under precisely
these conditions that the sorting and
retention system would be expected to
operate. Does collapse of damage segre-
gation, then, trigger aging? This failure is
accompanied by abnormal morphology
of mitochondria, which fragment and no
longer form a tubular network in old cells.
Whether aged yeast cells fail to segregate
protein aggregates asymmetrically re-
mains a crucial but unsettled question.
The concept of organelle-based reten-
tion of protein aggregates has been previ-
ously described for aggregates associ-
ated with vacuoles and nuclei (Spokoini
et al., 2012), though Zhou et al. could
not reproduce vacuolar tethering here.
The findings presented by Zhou et al.
nonetheless recalibrate the original pas-
sive model of aggregate retention to
include specific active organization of
protein aggregates, which is dependent
on mitochondria and factors, including
Fis1.
Several linked questions emerge
relating to this updated organelle-reten-
tion model. Which components mediate
tethering of protein aggregates to the ER
and mitochondria? Are older, potentially
less functional, mitochondria selected
for tethering targets in the first place,
allowing for simultaneous retention of
damaged organelles and aggregated
proteins in older cells? Which cellular sys-
tem ensures that mitochondria associ-
ated with aggregates do not leak into
daughter cells? Does the same retentioner Inc.mechanism operate in stem and progeni-
tor cells of the metazoa exhibiting asym-
metric damage distribution?
The organelle-retention model does not
easily accommodate or link to the alterna-
tive aggregate retention model centered
on actin cytoskeleton functions. Zhou
et al. show that inhibition of actin polymer-
ization by the drug Latrunculin A reduces
aggregate mobility but does not affect
aggregate association with mitochondria.
However, increasingly oxidized and less
functional mitochondria are preferentially
retained in mother cells, in a process
controlled by retrograde actin cable flow
(Higuchi et al., 2013). This points to further
cellular sorting systems that regulate and
impact cellular rejuvenation and lifespan.
Clarification of the role of the actin cyto-
skeleton in mitochondrial inheritance
might well end up dovetailing bothmodels
for cellular rejuvenation by mitochondrial
trash sorting.
ACKNOWLEDGMENTS
We thank L. Guilbride for thoughtful contributions
and critical editing and S. Miller for figure prepara-
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Mitochondria Tether Protein Trash to Rejuvenate Cellular Environments 

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Mitochondria Tether Protein Trash to Rejuvenate Cellular Environments

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