INSIGHTS | PERSPECTIVES
loid fibers—intrinsically disordered proteins
with low amino acid sequence complexity
(1, 4). They show that ATP substantially enhanced the solubility of the RNA-binding
protein fused in sarcoma (FUS), dissolving
its phase-separated droplets (see the figure).
This activity is not limited to FUS droplets
because ATP also dissolved liquid droplets of
other RNA-binding proteins and FUS amyloid fibers. Millimolar concentrations of ATP
decreased nucleation of FUS droplets and
fibers. These activities are consistent with
prevailing hydrotropic mechanisms of action.
The findings have interesting implications on
potential, previously unappreciated, roles of
ATP in biology and suggest a variety of additional studies.
Earlier work showed that ATP depletion
confined the mobility of different green fluorescent protein–tracer molecules in the cytoplasm of prokaryotic and eukaryotic cells.
Such metabolically inactive cells became solidlike (5). These effects were thought to be
due to a loss in the activity of ATP-dependent
machines. Observations by Patel et al. suggest that the hydrotrope activity of ATP also
may be acting to maintain a liquid cytoplasm
by keeping macromolecules soluble and preventing aggregation.
This activity may similarly play a role in
controlling the material properties of mem-
braneless organelles, such as nucleoli or
stress granules. Many of these organelles,
collectively called biomolecular condensates,
exhibit liquidlike behaviors, including fusion
and rapid exchange of material with the sur-
rounding solution (6). Evidence suggests,
however, that different condensates have dif-
ferent material properties, which can change
in response to signals (6). Previous work sug-
gested that ATP-dependent processes may
control these properties. For example, the
ATP-dependent disaggregating chaperone
heat shock protein 104 (Hsp104) increases
the rate of stress granule disassembly in yeast
(5). ATP-dependent enzymes such as RNA he-
licases are found in other condensates, such
as P-bodies in the cytoplasm (7). ATP-driven
active processes performed by these enzymes
are thought to control the physical proper-
ties of condensates and promote condensate
disassembly when required by the cell. Yet,
an unexplained quantitative feature of cellu-
lar biochemistry is that the cytoplasmic con-
centration of ATP is 10- to 100-fold above the
Michaelis constant (Km) of most ATP-utilizing
enzymes. Thus, an alternative, nonmutually
exclusive possibility is that the millimolar
concentration of ATP itself maintains dy-
namic, liquidlike condensates by promoting
constituent protein solubility and reducing
aggregation of phase-separating proteins.
Most healthy cell functions require that
proteins remain soluble at enormous intra-
cellular concentrations, without aggregating
into pathogenic deposits. Such aggregates are
found in many neurodegenerative diseases,
and current data suggest that some may arise
from aberrant phase separation. Intrinsically
disordered regions (IDRs), for example, can
mature into amyloid-like fibers, especially
when concentrated into a hydrogel or liquid
droplet in vitro (4, 8); such fibers can also be
observed in cells (9). The degree of fiber for-
mation is likely controlled physiologically to
match the needs of specific processes, and de-
fects in the balance between fiber-promoting
and fiber-inhibiting forces may lead to dis-
ease. For example, amyotrophic lateral scle-
rosis (ALS)–causing mutations in the IDRs
of FUS and heterogeneous nuclear ribonu-
cleoprotein A1 (hnRNPA1) promote amyloid
fiber formation (8, 10). Further, mutations
in valosin-containing protein (VCP/p97), an
adenosine triphosphatase that acts as a pro-
tein disaggregase, leave cells unable to clear
cytoplasmic stress granules, and patients
with VCP mutations present a multitude of
diseases, including familial ALS (11). Intrigu-
ingly, Drosophila models of VCP mutants
have decreased ATP amounts in the cell, and
mitochondrial dysfunction is found in many
neurodegenerative diseases (12). Further,
many neurodegenerative diseases are age
-related, and it is well established that cellu-
lar ATP concentrations decline with age. The
findings of Patel et al. suggest that ATP’s hy-
drotropic properties could act cooperatively
with machines such as VCP to decrease the
aggregation load on cells. Because hydrotrope
action is concentration-dependent, cells may
gradually lose their total disaggregase activity
as ATP abundance declines with age, making
them more prone to pathologic protein as-
sembly. This behavior would be exacerbated
in a mutagenic background of impaired VCP
activity or increased aggregation tendency of
proteins such as FUS or hnRNPA1.
Distinguishing the role that ATP plays in
driving active processes versus nonspecifically solubilizing proteins will remain a challenge for future researchers. In vivo work is
needed to confirm that modulating cellular
ATP concentrations results in the expected
phenotypes; increasing ATP should favor
condensate disassembly or accelerated molecular turnover, whereas ATP depletion
should favor more static, solidlike structures.
Further, researchers will need to correlate
changing ATP concentrations in aging cells
with quantitative changes in protein aggregation. Observations by Patel et al. suggest
a view of ATP as a molecule that not only
powers cellular machinery but also uses its
physical properties and high concentration
to control the material properties of biomolecular condensates and the cell interior. The
cell may exploit a natural hydrotrope to keep
itself in a functioning, dynamic state. j
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ATP controls solubility
Soluble, monomeric FUS More dynamic FUS droplets Less dynamic FUS droplets Insoluble FUS solids
Insoluble fber ATP FUS
Km values for ATP-driven cell processes typically lie between 10 and 500 mM, whereas hydrotrope activity requires 2 to 8 mM of ATP. Cells maintain
millimolar concentrations of ATP, perhaps to keep proteins (such as FUS) soluble by exploiting ATP as a biological hydrotrope.