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Supplementary Information accompanies the paper on www.nature.com/nature.
Acknowledgements We are grateful to C. Low for technical assistance; C. McArthur and S. Jiang
for cell sorting; R. Albert at Novartis Institutes for BioMedical Research for synthesis of
FTY720-P; and T. Okada, C. Allen and S. Watson for helpful discussions. M.M. is supported by
the Pfizer Postdoctoral Fellowship in Immunology and Rheumatology and the Rosalind Russell
Medical Research Center for Arthritis at University of California, San Francisco; J.G.C. is a
Packard fellow and an HHMI assistant investigator. This work was supported in part by grants
from the National Institutes of Health.
Competing interests statement The authors declare that they have no competing financial
interests.
Correspondence and requests for materials should be addressed to J.G.C. (cyster@itsa.ucsf.edu).
..............................................................
The mitochondrial calcium uniporter
is a highly selective ion channel
Yuriy Kirichok, Grigory Krapivinsky & David E. Clapham
Howard Hughes Medical Institute, Department of Cardiovascular Research,
Children’s Hospital, and Department of Neurobiology, Harvard Medical School,
320 Longwood Avenue, Boston, Massachusetts 02115, USA
.............................................................................................................................................................................
During intracellular Ca
21
signalling mitochondria accumulate
significant amounts of Ca
21
from the cytosol
1,2
. Mitochondrial
Ca
21
uptake controls the rate of energy production
1,3,4
, shapes
the amplitude and spatio-temporal patterns of intracellular Ca
21
signals
1,5–8
, and is instrumental to cell death
9,10
. This Ca
21
uptake
is undertaken by the mitochondrial Ca
21
uniporter (MCU)
located in the organelle’s inner membrane
11,12
. The uniporter
passes Ca
21
down the electrochemical gradient maintained
across this membrane without direct coupling to ATP hydrolysis
or transport of other ions
11
. Carriers are characterized by turn-
over numbers that are typically 1,000-fold lower than ion chan-
nels, and until now it has been unclear whether the MCU is a
carrier or a channel
13
. By patch-clamping the inner mitochon-
drial membrane, we identified a previously unknown Ca
21
-
selective ion channel sensitive to inhibitors of mitochondrial
Ca
21
uptake. Our data indicate that this unique channel binds
Ca
21
with extremely high affinity (dissociation constant #2 nM),
enabling high Ca
21
selectivity despite relatively low cytoplasmic
Ca
21
concentrations. The channel is inwardly rectifying, making
it especially effective for Ca
21
uptake into energized mitochon-
dria. Thus, we conclude that the properties of the current
mediated by this novel channel are those of the MCU.
Since its discovery in 1961 (ref. 14), mitochondrial Ca
2þ
uptake
has been measured in suspensions of isolated mitochondria. This
approach does not reliably control ion concentrations or voltage
gradients across the inner mitochondrial membrane, resulting in
inaccuracies in experimental results
11,12
. The patch-clamp method
solves these problems but its implementation in mitochondria is a
severe technical challenge. Here we patch-clamp single mitoplasts
(2–5-
m
m vesicles of inner mitochondrial membrane) isolated from
COS-7 cells (Fig. 1a, b), to measure directly whole-mitoplast and
single-channel Ca
2þ
currents.
In the whole-mitoplast configuration, voltage ramps from
2160 mV (potential across the inner membrane of energized
mitochondria) to þ80 mV elicited an inwardly rectifying current
that gradually increased as free Ca
2þ
concentration at the cytosolic
surface of the inner membrane ([Ca
2þ
]
c
) was varied from 20
m
Mto
100
m
M. These concentrations are comparable to [Ca
2þ
] in micro-
domains near endoplasmic/sarcoplasmic reticulum Ca
2þ
release or
plasma membrane Ca
2þ
channels, sensed by neighbouring mito-
chondria
1,2,15,16
. At 100
m
M [Ca
2þ
]
c
, the inwardly rectifying current
reached amplitudes of 20–30 pA (Fig. 1c). As mitoplasts are small,
this amplitude corresponds to a significant current density
(55 ^ 19 pA pF
21
at 2160 mV, n ¼ 4). To examine the level at
which the current saturates, [Ca
2þ
]
c
was elevated from 20
m
Mto
105 mM (Fig. 1c–e). Notably, the Ca
2þ
carrying capacity of the
current was enormous, reaching half-saturation at approximately
20 mM [Ca
2þ
]
c
. Elimination of Na
þ
in the bath did not reduce the
current, suggesting that Ca
2þ
was the primary charge carrier (not
shown). We refer to the mitoplast inwardly rectifying Ca
2þ
current
as I
MiCa
(for mitochondrial Ca
2þ
current).
I
MiCa
elicited by voltage steps rapidly inactivated to a plateau level
(Fig. 1f). The time-independent plateau was the major component
of the current with the transient component averaging 21 ^ 14% of
the net peak current (n ¼ 16). I
MiCa
amplitude was not noticeably
altered when pipette (intramitoplast) [Ca
2þ
]wasvariedfrom
,10 nM to about 10
m
M (no EGTA or EDTA). Thus, unlike many
Ca
2þ
-selective currents, I
MiCa
lacked Ca
2þ
-dependent inactivation
at energized mitochondrial potentials (approximately 2160 mV).
Similar to MCU-mediated mitochondrial Ca
2þ
uptake
11,17–19
,
Ruthenium 360 (Ru360) inhibited I
MiCa
in a dose-dependent
manner (half-maximal inhibitory concentration (IC
50
), 2 nM)
and was a more potent inhibitor than ruthenium red (RuR, IC
50
9 nM; Fig. 2).
RuR-sensitive (200 nM) I
MiCa
conducted Ca
2þ
and Sr
2þ
equally,
whereas Mg
2þ
was not measurably permeant (Fig. 3a). The relative
divalent ion conductance was Ca
2þ
< Sr
2þ
.. Mn
2þ
< Ba
2þ
(Fig. 3b), identical to the relative divalent ion permeability of the
MCU
13,20–22
. I
MiCa
currents carried by any of these four permeant
divalent ions were substantially reduced by low concentrations of
other permeant divalent ions, indicating competition for binding
at the selectivity site. The monovalent ions K
þ
and Na
þ
do not
contribute to I
MiCa
in the presence of [Ca
2þ
]
c
(Fig. 1d; see also
Supplementary Fig. 1). We refer to MiCa (mitochondrial calcium)
as the highly Ca
2þ
-selective conductance producing large current
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Group
densities in mitochondria, consistent with an ion channel.
Ion channel Ca
2þ
selectivity is the result of high-affinity Ca
2þ
binding to the permeation pathway, and removal of external Ca
2þ
enables these ion channels to conduct monovalent ions
23–26
. Indeed,
in low-divalent (5 mM EGTA/5 mM EDTA) solution, we recorded a
RuR- and Ru360-sensitive linear Na
þ
current (I
MiNa
). Interestingly,
I
MiNa
was relatively impermeant to K
þ
(Fig. 3c, top row). Increasing
[Ca
2þ
]
c
to 10 nM partially restored Ca
2þ
selectivity, whereas
100 nM led to complete inhibition of the Na
þ
conductance
(Fig. 3c, bottom left panel; estimated Ca
2þ
-binding equilibrium
constant #2 nM; see Methods). Although MiCa does not conduct
Mg
2þ
, in the absence of other divalent ions MiCa (I
MiNa
) binds
Mg
2þ
(Fig. 3c, bottom right panel) with an estimated binding
constant #250 nM (Methods). In 0.2 mM [Ca
2þ
]
c
,0.5mM
[Mg
2þ
]
c
reduced I
MiCa
to 41 ^ 8% of its conductance in nominally
Mg
2þ
-free solution (not shown). Thus, I
MiCa
will be reduced by
local cytosolic [Mg
2þ
]
c
, in agreement with data on the MCU
27,28
.
On the basis of its high sensitivity to RuR, Ru360 and Ca
2þ
,we
conclude that I
MiNa
is the monovalent current in divalent-free
conditions.
Single MiCa channel activity was recorded from inside-out
mitoplast patches. Under the ionic conditions used for the whole-
mitoplast configuration, 3–7 channels were active in each patch and
they closely mimicked the inward-rectifying I
MiCa
current–voltage
relationship (Fig. 4a). This density of single channels agrees with the
high I
MiCa
current density in whole mitoplasts. Single-channel
current amplitudes in 105 mM [Sr
2þ
]
c
(pipette) conditions were
similar to those in 105 mM [Ca
2þ
]
c
conditions; however, in agree-
ment with whole-mitoplast I
MiCa
selectivity, 105 mM [Mg
2þ
]
c
currents were not observed. RuR or Ru360 (200 nM; pipette)
rapidly blocked inward single-channel activity on stepping to
2180 mV (Fig. 4e). Block by 200 nM RuR and Ru360 was relieved
on returning to potentials above 0 mV (not shown). Thus, RuR and
Ru360 were ineffective in blocking i
MiCa
outward current (where i
indicates unitary current).
In symmetrical 105 mM [Ca
2þ
] conditions, outward single-
channel openings at positive voltages (always correlated with inward
openings at negative voltages) rapidly flickered between open and
closed states (Fig. 4a). Substitution of bath (matrix) Ca
2þ
by Mg
2þ
abolished these outward channels (Fig. 4b), whereas Sr
2þ
-mediated
outward currents were comparable to those in Ca
2þ
. In symmetrical
150 mM Na-gluconate low-divalent solution, single i
MiNa
Na
þ
channel currents had about tenfold higher conductance than
i
MiCa
, were selective for Na
þ
over K
þ
, exhibited multiple conduc-
tance states, and were blocked by 200 nM RuR. The rapid flickering
of outward single-channel currents observed in 105 mM Ca
2þ
were
not observed for the outward monovalent current. This suggests
that open channels are transiently blocked by outward-going Ca
2þ
Figure 1 Ca
2þ
current through the inner mitochondrial membrane. a, COS-7 cell
transfected with mitochondrial-targeted YFP. b, Transmitted and fluorescent images of a
mito-YFP-labelled mitoplast. Scale bar: 3
m
m. c, I
MiCa
elicited by voltage ramps. Negative
current flows from the cytoplasmic face of the inner mitochondrial membrane (bath) to the
interior of the mitoplast. Bath ¼ 0 mV. Inside mitoplast (pipette), Cs-gluconate solution;
bath, HEPES-Tris solution. d, I
MiCa
increases with bath [Ca
2þ
]
c
. Bath, Na-gluconate
solution. e, Peak amplitude of I
MiCa
at 2160 mV at varying [Ca
2þ
]
c
. Currents were
normalized to peak I
MiCa
in 105 mM [Ca
2þ
]
c
and fitted by I ¼ I
max
=ð1 þ
ðK
0:5
=½Ca
2þ
Þ
h
Þ; n ¼ 5. f, I
MiCa
in response to voltage steps (5 mM [Ca
2þ
]
c
Na-
gluconate solution); DV ¼ 20 mV.
letters to nature
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© 2004
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Group
ions. Indeed, the amplitude of ensemble-averaged Ca
2þ
currents
from multi-channel patches was about nine times less in the out-
ward than in the inward direction (Fig. 4c).
To determine single-channel kinetics from patches containing
only one active channel (Fig. 4d), smaller areas of membrane
were patched by using greatly reduced pipette tip diameters. In
symmetrical 105 mM CaCl
2
solution, the activity of single Ca
2þ
currents was time-independent at all voltages between 2200 and
þ140 mV. Single MiCa channels had multiple subconductance
states between 2.6 pS and 5.2 pS (2160 mV). Most importantly,
the open probability (P
open
) of the channel was about 99% at
2200 mV and declined to approximately 11% at 280 mV. This
dependence of P
open
on membrane potential accounts for much of
the inward rectification of the macroscopic I
MiCa
current over this
range of potentials (Supplementary Fig. 2).
In summary, a unique Ca
2þ
-selective channel of the inner
mitochondrial membrane accounts for the properties of the
MCU. MiCa is the only known functional intracellular Ca
2þ
-
selective channel (inositol-1,4,5-trisphosphate and ryanodine
Figure 2 RuR and Ru360 sensitivity. a, RuR (2–200 nM) inhibited I
MiCa
which did not fully
recover by 7 min after washout (red trace). The pipette (intramitoplast) contained Na-
gluconate solution whereas the HEPES-Tris bath solution contained 5 mM Ca
2þ
. b, Dose
response of I
MiCa
to RuR (n ¼ 5). c, I
MiCa
inhibition by the MCU blocker Ru360. I
MiCa
recovered 4 min after washout (red trace). The pipette was filled with Na-gluconate
solution (no EGTA/EDTA). d, Estimated dose–response curve of I
MiCa
in response to
Ru360. The time courses of inhibition were fitted by single exponential functions with
t ¼ 5 ^ 2s(n ¼ 4) at 2 nM and 270 ^ 160 ms (n ¼ 4) at 50 nM. Steady state
inhibition is estimated from fits for 2 and 5 nM.
Figure 3 I
MiCa
selectivity. a, I
MiCa
in 5 mM Ca
2þ
,Sr
2þ
,Mn
2þ
,Ba
2þ
(blue traces) or
Mg
2þ
(black traces). Recordings are from the same mitoplast in HEPES-Tris-based
solution, washed between applications with 1 mM EGTA/EDTA solution. Pipettes
contained Na-gluconate solution. b, Relative conductance of I
MiCa
to divalent cations.
Averaged peak current densities at 2160 mV (n ¼ 5; ^s.d.). c, I
MiCa
conducts Na
þ
in
the absence of divalent ions (I
MiNa
). Top left panel: I
MiNa
in Na-gluconate (black trace) and
K-gluconate (red trace) low-divalent (LD) solution. Top right panel: I
MiNa
Na
þ
(LD) current
was inhibited by RuR and Ru360. Bottom left panel: [Ca
2þ
]
c
at 10 nM and 100 nM
inhibited I
MiNa
. Bottom right panel: [Mg
2þ
]
c
at 1
m
M inhibited I
MiNa
.
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Group
receptors are nonselective, but are Ca
2þ
-permeant). MiCa is Ca
2þ
-
selective even in low cytoplasmic [Ca
2þ
], it conducts Ca
2þ
prefer-
entially into mitochondria, has relative divalent conductances
Ca
2þ
< Sr
2þ
.. Mn
2þ
< Ba
2þ
; is blocked by nanomolar concen-
trations of RuR and Ru360, and can provide sufficient current
densities to explain mitochondrial uptake by the MCU.
There is one marked difference between our data and Ca
2þ
flux
studies using suspensions of isolated mitochondria. Here, in
105 mM [Ca
2þ
]
c
, I
MiCa
is close to saturation with a maximum
ion flux through the single MiCa molecule of approximately 5 £ 10
6
Ca
2þ
s
21
(half-activation constant, K
0.5
¼ 19 mM), significantly
higher than the approximately 2 £ 10
4
Ca
2þ
s
21
and K
0.5
of about
10
m
M estimated for MCU from suspensions of isolated mitochon-
dria
11,13
. This difference in flux estimates stems from the fact that in
mitochondrial suspensions, the rapid dissipation of the mitochon-
drial potential during Ca
2þ
entry
11,12
would appear to cause satura-
tion of Ca
2þ
flux at much lower [Ca
2þ
]
c
. At [Ca
2þ
]
c
above 100
m
M,
I
MiCa
current densities are so large that the potential across the inner
membrane can only be maintained by voltage clamp. On the basis of
an I
MiCa
density of 1,000–2,000 pA pF
21
, .90% open probability,
and 0.5–1 pA single-channel amplitude at ½Ca
2þ
c
¼ 105 mM and
V
m
¼ 2160 mV, MiCa channel density in the inner mitochondrial
membrane is approximately 10–40 channels per
m
m
2
, slightly lower
than voltage-gated Ca
2þ
channel density (about 100 per
m
m
2
) (ref.
29) in excitable cell membranes. Nonetheless, at micromolar con-
centrations of [Ca
2þ
]
c
, MiCa channels provide current densities
comparable to those of voltage-gated Ca
2þ
channels at millimolar
Ca
2þ
concentrations. This feat is accomplished by the large electro-
chemical gradient for Ca
2þ
across the inner membrane of energized
mitochondria and the unusually high probability of MiCa being in
the open state. Ca
2þ
-dependent inactivation, a negative feedback
mechanism for Ca
2þ
-permeant channels
29
, is surprisingly absent for
MiCa at normal potentials for energized mitochondria.
Na
þ
current through MiCa appears only at extremely low
concentrations of Ca
2þ
, suggesting that binding sites inside the
pore have an exceptionally high affinity (dissociation constant
#2 nM) for Ca
2þ
. This property guarantees its high selectivity for
cytosolic Ca
2þ
where monovalent cations outnumber Ca
2þ
ions by
10
3
–10
6
-fold. MiCa is impermeant to Mg
2þ
and K
þ
, thus prevent-
ing mitochondrial depolarization by these abundant ions. In short
the high Ca
2þ
selectivity of MiCa results in mitochondrial Ca
2þ
accumulation with the least dissipation of mitochondrial potential.
On the basis of the properties demonstrated in our experiments, we
conclude that the mitochondrial Ca
2þ
uniporter is the Ca
2þ
-
selective ion channel MiCa. A
Methods
Preparation of the mitoplasts
COS-7 cells were homogenized by a Teflon pestle in a glass grinder with 250 mM sucrose,
5 mM HEPES and 1 mM EGTA (pH 7.2 with KOH). Mitochondria were isolated from the
lysed COS-7 cell by differential centrifugation in the same solution
30
. To obtain mitoplasts,
mitochondria were subjected to osmotic shock for 5 min in hypotonic solution containing
5 mM sucrose, 5 mM HEPES and 1 mM EGTA (pH 7.2 with KOH). Mitoplasts were
sedimented from this solution by centrifugation at 3,920g
max
for 5 min and subsequently
re-suspended in 750 mM KCl, 100 mM HEPES and 1 mM EGTA (pH 7.2 with KOH).
Mitoplast isolation was carried out at 2 8C and mitoplasts stored on ice. Just before
experiments, 1–3
m
l of the mitoplast suspension was added to the experimental solution
containing 150 mM Na-gluconate, 10 mM HEPES, 1 mM EGTA and 1 mM EDTA (pH 7.2
with NaOH). Isolated mitoplasts were transparent vesicles with one to several black spots,
presumably formed by the remnants of the outer membrane in contact sites between two
mitochondrial membranes. In some experiments, mitoplasts were isolated from COS-7
cells transfected with yellow fluorescent protein (YFP) targeted to the inner membrane of
mitochondria (BD Clontech) to confirm the mitochondrial origin of the vesicles.
Mitoplasts of 2–5
m
M diameter (membrane capacitance, C
m
¼ 0.35–1 pF) were used for
patch-clamp experiments.
Whole-mitoplast recordings
After formation of a GQ seal between the patch-clamp pipette and inner mitochondrial
membrane, capacitance transients were completely compensated. Voltage steps of 200–
500 mV and 3–300 ms were then applied to rupture the membrane and obtain the whole-
mitoplast configuration as monitored by reappearance of capacitance transients and
increase in baseline noise. In some experiments BSA (0.05%) was added to the bath
solution to increase the survival of mitoplasts. Elevation of pipette solution tonicity (£1.5
bath solution) significantly reduced the access resistance associated with the mitoplast
shrinkage after break-in. The ionic compositions of pipette and bath solutions were
chosen to reduce potential contamination by K
þ
and Cl
2
currents, especially in the inward
Figure 4 Single i
MiCa
Ca
2þ
channels from inner mitochondrial membrane inside-out
patches (105 mM CaCl
2
solution at the cytoplasmic surface (pipette)). a, Multi-channel
patch currents. The bath contained 150 mM Na-gluconate, 1 mM EGTA/EDTA (free Ca
2þ
concentration at the matrix surface of the inner membrane ([Ca
2þ
]
m
) ,10 nM) (red trace)
or 105 mM [Ca
2þ
]
m
(black trace). b, i
MiCa
in 105 mM [Ca
2þ
]
m
(black trace) or 105 mM
[Mg
2þ
]
m
(red trace). c, Ensemble averages of 45 multi-channel responses. Voltage ramp:
2160 mV to þ160 mV. d, Rare single i
MiCa
. [Ca
2þ
]
m
and [Ca
2þ
]
c
were both 105 mM.
Note the fast open/close kinetics of the attenuated outward current. e, [RuR]
c
or [Ru360]
c
in the pipette inhibited i
MiCa
at 200 nM. Red traces indicate ensemble averages of 20
multi-channel responses.
letters to nature
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Group
direction. Three different pipette solutions were used: Cs-gluconate solution (150 mM
CsOH, 5 mM CsCl, 135 mM sucrose, 10 mM HEPES, 1.5 mM EGTA and 1.5 mM EDTA
(pH 7.2 with
D-gluconic acid)); Na-gluconate solution (150 mM Na-gluconate, 5 mM
NaCl, 135 mM sucrose, 10 mM HEPES, 1.5 mM EGTA and 1.5 mM EDTA (pH 7.2 with
NaOH)); Na-gluconate solution without Ca
2þ
buffer (150 mM Na-gluconate, 5 mM NaCl,
135 mM sucrose, 10 mM HEPES (pH 7.2 with NaOH)).
Signals were recorded using an Axopatch 200B patch-clamp amplifier, filtered at 5 kHz,
and sampled at 10 kHz. Capacitance of the whole-mitoplast membrane ranged from 0.35
to 1 pF. Access resistances, usually 30–60 MQ, were compensated by about 70%. Bath Ca
2þ
concentration was varied from 20
m
M to 5 mM by addition of the corresponding amount
of 1 M CaCl
2
stock solution into one of two solutions: Na-gluconate solution (150 mM
Na-gluconate, 10 mM HEPES (pH 7.2 with NaOH)) or HEPES-Tris solution (205 mM
HEPES (pH 7.2 with Trisma base)). Bath solution with 105 mM [Ca
2þ
]
c
(105 mM CaCl
2
solution) contained 105 mM CaCl
2
, 10 mM HEPES (pH 7.2 with Trisma base or NaOH).
This solution was dissolved with Na-gluconate solution or HEPES-Tris solution to obtain
26 mM [Ca
2þ
]
c
. Bath solutions with 5 mM of Sr
2þ
,Ba
2þ
,Mn
2þ
or Mg
2þ
were obtained by
addition of 1 M stock solution of the chloride salt into HEPES-Tris solution. HEPES-K
bath solution (KOH 75 mM (pH 7.2 with about 210 mM HEPES)); HEPES-Na bath
solution (NaOH 75 mM (pH 7.2 with about 210 mM HEPES)); Na-gluconate and K-
gluconate low-divalent solutions (150 mM Na(K)-gluconate, 10 mM HEPES, 5 mM
EGTA, 5 mM EDTA (pH 7.2 with NaOH)). CaCl
2
or MgCl
2
was added to Na-gluconate
low-divalent (divalent-free) solution in the amounts calculated by the WinMAXC v2.05
program (C. Patton, Stanford University) to obtain free Ca
2þ
and Mg
2þ
concentrations.
Osmolarity was approximately 280 mmol kg
21
. Statistical data was calculated as the
mean ^ s.d.
Single-channel recordings
All single-channel recordings were made in the inside-out configuration of the patch-
clamp technique. Patches were excised from the inner mitochondrial membrane in a bath
solution containing 150 mM Na-gluconate, 10 mM HEPES, 1 mM EGTA and 1 mM EDTA
(pH 7.2 with NaOH). Pipettes were filled with 105 mM CaCl
2
solution. A total of 105 mM
CaCl
2
solution was also used as the bath solution for single channels and whole-mitoplast
recordings. Signals were filtered at 1 kHz and sampled at 5 kHz. In Supplementary Fig. 2,
traces were filtered at 200 Hz before amplitude analysis.
Estimation of Ca
21
- and Mg
21
-binding constants
Ca
2þ
buffering is unreliable below 10 nM. To estimate the MiNa Ca
2þ
-binding constant,
we assumed that I
MiNa
in low-divalent solution was #I
MiNa
at theoretical 0 [Ca
2þ
]
c
. Given
that I
MiNa
at 10 nM [Ca
2þ
]
c
was about eightfold less than I
MiNa
in low-divalent solution,
the Ca
2þ
-binding equilibrium constant calculated from the Hill equation was #2nM
assuming a Hill coefficient of 1. Similarly, the MiNa-binding constant for Mg
2þ
was
estimated to be #250 nM.
Received 20 August; accepted 18 November 2003; doi:10.1038/nature02246.
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Supplementary Information accompanies the paper on www.nature.com/nature.
Acknowledgements We would like to thank J. Borecky for advice on the whole-mitoplast
configuration; L. DeFelice, H. Xu, B. Desai, V. Sandler and P. Smith for discussions; and S. Gapon
and Y. Manasian for technical assistance.
Competing interests statement The authors declare that they have no competing financial
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..............................................................
Two mitotic kinesins cooperate
to drive sister chromatid
separation during anaphase
Gregory C. Rogers
1
, Stephen L. Rogers
2
, Tamara A. Schwimmer
1
,
Stephanie C. Ems-McClung
3
, Claire E. Walczak
3
, Ronald D. Vale
2
,
Jonathan M. Scholey
4
& David J. Sharp
1
1
Department of Physiology and Biophysics, Albert Einstein College of Medicine,
Bronx, New York 10461, USA
2
The Howard Hughes Medical Institute and Department of Cellular and
Molecular Pharmacology, University of California, San Francisco, California
94143, USA
3
Medical Sciences Program, Indiana University, Bloomington, Indiana 47405,
USA
4
Center for Genetics and Development and Section of Molecular and Cellular
Biology, University of California, Davis, California 95616, USA
.............................................................................................................................................................................
During anaphase identical sister chromatids separate and move
towards opposite poles of the mitotic spindle
1,2
. In the spindle,
kinetochore microtubules
3
have their plus ends embedded in the
kinetochore and their minus ends at the spindle pole. Two models
have been proposed to account for the movement of chromatids
during anaphase. In the ‘Pac-Man’ model, kinetochores induce
the depolymerization of kinetochore microtubules at their plus
ends, which allows chromatids to move towards the pole by
‘chewing up’ microtubule tracks
4,5
. In the ‘poleward flux’ model,
kinetochores anchor kinetochore microtubules and chromatids
are pulled towards the poles through the depolymerization of
kinetochore microtubules at the minus ends
6
. Here, we show that
two functionally distinct microtubule-destabilizing KinI kinesin
letters to nature
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