Post on 18-Nov-2020
Recent Advances in Vitamin K Metabolism
3rd International Conference on Nutrition and Food Science(Nutritional Science-2014)Track 1: Nutrition and Basic Science14:10-14:30, September 23, 2014Committee Room 1-2Palacio de Congresos de Valencia, Spain
Kobe Pharmaceutical UniversityToshio Okano
Dept. Hygienic SciencesKobe Pharmaceutical Univ.
O
O
O
O
Naturally occurring vitamin K
Plant origin(Major dietary
source)
PK (phylloquinone: vitamin K1)
Bacterial origin(Minor dietary
MK-4 (menaquinone-4: vitamin K2)MK-7 O
O
Plasma vitamin K concentrations of healthy Japanese women
phylloquinone (ng/mL) MK-4 (ng/mL)1.74±1.29 (0.13〜8.83) 0.10±0.19 (n.d.〜1.44)
(Minor dietary
source) MK-7 (menaquinone-7)
Tsugawa N, Okano T, et al, Am J Clin Nutr, 2006;83:380-386.
Tissue concentrations of animals and humansMK-4>> phylloquinone
Amounts of dietary intake and plasma concentrations phylloquinone>> MK-4
Is phylloquinone converted into MK-4 in the body ?
The aim of our study is to examine the above issues in animals and humans.
What is the physiological significance of this conversion and what functions dose MK-4 have, and can we develop MK-4
analogues for clinical use ? Suhara Y, Okano T, et al, J Med Chem, 2011;54:4269-4273
Suhara Y, Okano T, et al, J Med Chem, 2011;54:4918-4922
Suhara Y, Okano T, et al, J Med Chem, 2012;55;1553-1558
Tissue concentrations of PK and MK-4 in mice fed a conventional laboratory chow diet
0
5
10
15
20
25
30
35
40
PK C
onc.(pm
ol/g)
PKPKPKPK
PKPKPKPK epoxideepoxideepoxideepoxide
0
200
400
600
800
1000
1200
1400
1600
1800
2000
MK-4 C
onc.(pm
ol/g)
MK-4
MK-4 epoxide
×100
Okano T. et al., JBC 2008;283:11270-11279
LC-MS and NMR
PK
Authentic vitamin KsMK-4
MK-4 fraction
Inte
ns
ity,
cp
s
MK-4
Identification of MK-4 from brain of mice by LC-MS/MS
MK-4
MK-4 epoxide
PK epoxide
Time,min
PKPK
epoxide
Time,min
PKMK-4
epoxidePK
epoxide
Inte
ns
ity,
cp
s
MK-4 epoxide PK
PK epoxide
H-5,88.06 H-6,7
7.67
H-2’,6’,10’,14’5.03
H-1’3.45
CH3((((C-2))))2.17
Authentic MK-4
Identification of MK-4 from brain of mice by1H NMR spectrometryO
O
1 2
345
6
7
8
2’ 6’ 10’ 14’
1’
7.67 5.03
012345678δδδδ((((ppm))))
H-5,88.06
H-6,77.68
H-2’,6’,10’,14’5.08
H-1’3.35
MK-4 fraction CH3((((C-2))))2.17
1 w 24 hr
PK-d7
12 hr
C57BL/6J mice7-weeks old
Fast Sacrifice
0
Oral administration
Cerebra
Experimental design
Experiment
7-weeks oldn=200
Analyses by LC-MS/MS and 2H NMR
Isolation and purification of MK-4-d7
O
O 3
PK-d7
D
D
D
D
CD3
O
O 3
MK-4 -d7
D
D
D
D
CD3????
cps)
4.5e4
3.0e4Inten
sity(
cps)
0
Authentic MK-4-d7
MK-4-d7 fraction
194.3452.5
Identification of MK-4-d7 from brain of mice byLC-MS/MS
3.5e4
1.5e4
((((m/z))))
Inten
sity(
cps
100 200 300 400 50000
O
O
CH3
CD3
D
D
D
D
m/z:194
194.4452.6
+ H
2H-5,8 C2H
Authentic MK-4-d7
O
O
CD3
D
D
D
D
1 2
345
67
8MK-4-d7
Identification of MK-4-d7 from brain of mice by2H NMR spectrometry
H-5,88.10
2H-6,77.70
C2H3
2.14
012345678δδδδ((((ppm))))
2H-6,77.68
2H-5,88.08
MK-4-d7fractionC2H3
2.14
Not only PK-d7 but also MK-4-d9 and MK-7-d7 are converted into MK-4-d7 and accumulate in brain of mice
O
O 3
D
D
D
D
CD3PK-d7
O
O 3
D
D
D
D
CD3
D D
MK-4-d9 MK-7-d7O
O 6
D
D
D
D
CD3
24 hr12 hr
Fast Sacrifice
0
C57BL/6J ♀mice
8 weeks old
O
O 3
D
D
D
D
CD3 MK-4-d7
Single peroral administration (10 µg/kg)0
20
40
60
80
100
120
vehicle PK-d7 MK-4-d9
pmol/
gof b
rain
MK-7-d7
MK-4MK-4 epoxide
MK-4-d7
MK-4-d7 epoxideEndogenous MK-4
Phylloqunone is converted into MK-4 via integral side-chain removal
Unsaturation of side chain
O
O 3
O
O 3
O
O 3
O
O 3
DDDD
MK-4-d9CD3
D
D
D
D
D D
CD3
D
D
D
D
D D
K1-d9
D DD D
CD3
D
D
D
D
CD3
D
D
D
D
OO Exchange of OO
D DD D
MK-4-d7CD3
D
D
D
D
CD3
D
D
D
D
CD3
D
D
D
D
CD3
D
D
D
D
O
O
O
OPhytyl side chain
Geranylgeranylside chain
33
3
PP
PO O
O-
OP :
PO O-
O-
OP
PO O
O-
OP :
PO O-
O-
OPK3
Exchange of
side chain
O
O 3
O
O 3
K vitamins are converted into MK-4 and accumulate in
tissuesWhere does this conversion take
place?Following four routes for the Following four routes for the conversion of PK or K3 into MK-4,
1.Oral route2.Enteral route3.Intravenous route4.Intra-cerebroventricular
routewere examined in mice.
1 w 24 hr
K1-d7, K3-d8
12 hrC57BL/6J mice
7 weeks old
Fast Sacrifice
0
Oral(1), enteral(2), intravenous(3) or intra-cerebroventricular(4) dose
Cerebra
LC-APCI//MS
Experimental design
A single dose of 10 μmol/kg BW for (1), (2), (3) At 0.1 μmol/Kg BW for (4)
O
O 3
K1-d7
D
D
D
D
CD3
O
O 3
MK-4-d7
D
D
D
D
CD3
O
O K3-d8
D
D
D
D
CD3
D
Concentrations of MK-4 in cerebra of mice
0
50
100
150
200
Co
nc
en
tra
tio
n
(( ((p
mo
l/g)) ))
Vehicle K3-d8PK-d Vehicle K3-d8PK-d7
0
100
200
300
MK-4 MK-4 epo MK-4-d7 MK-4-d7 epo
Endogenous MK-4 Converted MK-4
oral entero
Vehicle K3-d8PK-d7Vehicle K3-d8PK-d7
Vehicle K3-d8PK-d7
0
50
100
150
200
250
Co
nc
en
tra
tio
n
(( ((p
mo
l/g)) ))
Vehicle K3-d8PK-d70
50
100
150
200
250intravenous Intra-cerebroventricular
Okano T, et al, J Biol Chem, 2008; 283:11270-11279
Four Sites Cannulation experiments
Bile duct Portal vein
Inferior vena cava
Thoracic Lymph duct
heartheartheartheart
liver
serum (IVC)
Lymph(TLD)
Po K1-d7
0 1 62 3 4 5 (h)
sacrificeWistar rat
Bile duct Portal vein
intestineintestineintestineintestine
Bile (BD) Serum (PV)
6080100181.10525
[M+H]+
Authentic MD-d8 173.10173.08173.06173.04173.02173.00020406080100
173.05982[M+H]+
Authentic MD
(m/z )Relative Abundance (%
)ODD CD3
OORelative Abundance (%
)0102030405060
0 1 2 3 4 5 6Concentration(pmol/g) bileSerum from IVCSerum from PVLymph
0.030.040.05 MK-4-d7
K1-d7
oncentration(pmol/g)
high resolution mass spectrometry
(HR-MS)
bileSerum from IVCSerum from PVLymph
Concentrations of K1-d7, MK-4-d7 and MD-d7
180.06 180.08 180.10 180.12 180.14 180.16020406080100
180.10172[M+H]+
181.16181.14181.12181.10181.08181.060204060[M+H]+
MD fraction from lymphOODDD D CD3OD D 3D
(m/z )(m/z )
Relative Abundance (%)
Relative Abundance (%)00.010.02
0 1 2 3 4 5 6Time after administration (h)00.40.81.2
0 1 2 3 4 5 6Time after administration (h)Concentration (pmol/mL) MD-d7Concentration
bileSerum from IVCSerum from PVLymph MD-d7
400
con
cen
tra
tio
n(p
mo
l/m
L)
600
4 c
on
cen
tra
tio
n(p
mo
l/m
L)
6
8
10
12
18
24
MD
co
nce
ntr
ati
on
(pm
ol/
mL)
K1 MK-4 MD
Time course changes in serum concentrations of K1, MK-4 and MD in humans orally given K1 capsules (40 mg)
(n=5) (n=5) (n=5)
200
0 1 2 3 4 5 6
K1
con
cen
tra
tio
n
0 1 2 3 4 5 6
MK
-4 c
on
cen
tra
tio
n(p
mo
l/m
L
0
2
4
0
6
12
0 1 2 3 4 5 6
MD
co
nce
ntr
ati
on
(pm
ol/
mL
Time after administration (h)Time after administration (h)Time after administration (h)
0
GGPP
MK-4-d2
Route 1
Conversion pathway of MD-d3 to MK-4-d3 demonstrated by HR-MS and
1H NMR analyses (Route 2)
MD-d
GGPPMK-4-d3
Route 2
MD-d3
Hirota Y, Okano T, et al, J Biol Chem, 2013; 288:33071-33080
K1
Removal of side chainSide-chain cleavage
enzyme
MD (K3)(quinone form))
Lymph and blood stream
(intestine)
MD(K3) is a catabolic product of oral phylloquinone (K1) in the intestine and
a circulating precursor of tissue MK-4 (K2) in mammals
Prenylation enzyme
Geranylgeranyl pyrophosphate
MK-4 (K2)
Reductase (tissues)
℗-℗℗-℗:
Hirota Y, Okano T, et al, J Biol Chem, 2013; 288:33071-33080
MD (K3)(hydroquinone form))
New paradigm of the metabolic activation of
vitamin K in brain and bone
Biological actions
O
O 3
O
phylloquinone(PK)
GGCX, SXR/PXR, PKA/PKC
brain
Menaquinone-4((((MK-4))))
O
O 3
active metabolite
O
O
Foods
OR n-1
R:
menaquinones(MKn)
Menadione(MD)
bone
O
O 3
GGCX, SXR/PXR, PKA/PKC
Biological actions
O ?
What is the enzyme(s) involved in MK-4
biosynthesis in mammals ?
phylloquinone(PK)
O
OMenaquinone-4((((MK-4))))
O
O 3
foods
OR n-1
R:
Enzyme(s)
active metabolite
Menadione(MD)
menaquinones(MKn)
Biosynthesis of menaquinones in Escherichia. coli
menB
menF
menEmenCmenD
COO-
OHO COO-
Chorismate
O-Succinylbenzoyl-CoA 1,4-Dihydroxy-2-napthol-CoA
O
-OOCOH
COO-
Isochorismate
OH
OH
S-CoA
O
COO-
S-CoA
O
O
n
P2O73- P2O7
3-
nH
OH
OH
menG
yfbB
1,4-Dihydroxy-2-napthanoate Demethlmenaquinone
hydroquinone((((DMK-n))))
Menaquinone-nhydroquinone((((MK-n))))
Menaquinone-n
((((MK-n))))
O-
OH
OH OCO2
nP2O7
n
OH
OH
H
O
O n
menA
Alignment of the amino acid sequence of Ubia(E.coli), COQ2(Homo-sapiens), Men A(E.coli)
and UBIAD1(Homo-sapiens) ubiA_[Escherichia_coli] 1 ------------------------------------------------------------
COQ2_[Homo_sapiens] 1 MLGSRAAGFARGLRAVALAWLPGWRGRSFALARAAGAPHGGDLQPPACPEPRGRQLSLSA
_menA_[Escherichia_coli] 1 ------------------------------------------------------MTEQQI
UBIAD1_[Homo_sapiens] 1 ----------------------------MAASQVLGEKINILSGETVKAGDRDPLGNDCP
ubiA_[Escherichia_coli] 1 -MEWSLTQNKLLAFHRLMRTDKPIGALLLLWPTLWALWVATPG--VPQLWILAVFVAGVW
COQ2_[Homo_sapiens] 61 AAVVDSAPRPLQPYLRLMRLDKPIGTWLLYLPCTWSIGLAAEPGCFPDWYMLSLFGTGAI
_menA_[Escherichia_coli] 7 SRTQAWLESLRPKTLPLAFAAIIVGTALAWWQGHFDPLVALLALITAGLLQILSNLANDY
UBIAD1_[Homo_sapiens] 33 EQDRLPQRSWRQKCASYVLALRPWSFSASLTPVALGSALAYRSHGVLDPRLLVGCAVAVL
ubiA_[Escherichia_coli] 58 LMRAAGCVVNDYADRKFDGHVKRTANRPLPSGAVTEKEARALFVVLVLISFLLVLTLNTM
COQ2_[Homo_sapiens] 121 LMRGAGCTINDMWDQDYDKKVTRTANRPIAAGDISTFQSFVFLGGQLTLALGVLLCLNYY
_menA_[Escherichia_coli] 67 GDAVKGSDKPDRIGPLRGMQKGVITQQEMKRALIITVVLICLSGLALVAVACHTLADFVG
UBIAD1_[Homo_sapiens] 93 AVHGAGNLVNTYYDFSKGIDHKKSDDRTLVDRILEPQDVVRFGVFLYTLGCVCAACLYYL
ubiA_[Escherichia_coli] 118 T------ILLSIAALALAWVYPFMKR---YTHLPQVVLGAAFGWSIPMAFAAVS-ESVPL ubiA_[Escherichia_coli] 118 T------ILLSIAALALAWVYPFMKR---YTHLPQVVLGAAFGWSIPMAFAAVS-ESVPL
COQ2_[Homo_sapiens] 181 S------IALGAGSLLLVITYPLMKR---ISYWPQLALGLTFNWGALLGWSAIKGSCDPS
_menA_[Escherichia_coli] 127 F------LILGGLSIIAAITYTVGNRPYGYIGLGDISVLVFFGWLSVMGSWYLQAHTLIP
UBIAD1_[Homo_sapiens] 153 SPLKLEHLALIYFGGLSGSFLYTGGIGFKYVALGDLIILITFGPLAVMFAYAIQVGSLAI
ubiA_[Escherichia_coli] 168 SCWLMFLANILWAVAYDTQYAMVDRDDDVKIGIKSTAILFGQY-DKLIIGILQIGVLALM
COQ2_[Homo_sapiens] 232 VCLPLYFSGVMWTLIYDTIYAHQDKRDDVLIGLKSTALRFGEN-TKPWLSGFSVAMLGAL
_menA_[Escherichia_coli] 181 ALILPATACGLLATAVLNINNLRDINSDRENGKNTLVVRLGEVNARRYHACLLMGSLVCL
UBIAD1_[Homo_sapiens] 213 FPLVYAIPLALSTEAILHSNNTRDMESDREAGIVTLAILIGPT--FSYILYNTLLFLPYL
ubiA_[Escherichia_coli] 227 AIIGELNGLGWGYYWSILVAGALFVYQQKLIANREREACFKAFMNNNYVGLVLFLGLAMS
COQ2_[Homo_sapiens] 291 SLVGVNSGQTAPYYAALGAVGAHLTHQIYTLDIHRPEDCWNKFISNRTLGLIVFLGIVLG
_menA_[Escherichia_coli] 241 ALFNLFSLHSLWGWLFLLAAPLLVKQARYVMREMDPVAMRPMLERTVKGALLTNLLFVLG
UBIAD1_[Homo_sapiens] 271 VFSILATHCTISLALPLLTIPMAFSLERQFRSQAFNKLPQRTAKLNLLLGLFYVFGIILA
ubiA_[Escherichia_coli] 287 -YWHF----------------
COQ2_[Homo_sapiens] 351 NLWKEKKTDKTKKGIENKIEN
_menA_[Escherichia_coli] 301 IFLSQWAA-------------
UBIAD1_[Homo_sapiens] 331 PAGSLPKI-------------
Search for the enzyme(s) responsible for the MK-4 synthesis
E.coliOH
OH
O-
O
1,4-Dihydroxy-2-naphthoate
((((DHNA))))
nH
OH
OH
Demethylmenaquinone-n
hydroquinone((((DMK-n))))
MenA
Menaquinone-n
((((MK-n))))
MenG
H
O
On
nH
Menaquinone-n biosynthesis in E.coli
MenA human homologs
Chromosome 1 11.25 MMenA Prenylation domain 311 a.a
O
O 3
MK-4O
O
+3
P P
GGPPMD
UBIAD1 orCOQ2 ?
338 a.a
384 a.a
Chromosome 1 11.25 M
Chromosome 4 84.40 M
MenA
UbiA prenyltransferasedomain containing 1
((((UBIAD1))))
CoenzymeQ2 homolog,prenyltransferase
((((COQ2))))
Prenylation domain
Prenylation domain
311 a.a
Prenylation domain
Co
nce
ntr
ati
on
(p
mo
l/m
g p
rote
in)
Conversion of K3-d8 or MK-4-d12 to MK-4-d7 in siControl-, siUBIAD1- or siCOQ2-transfected MG-63 cells
Co
nce
ntr
ati
on
(p
mo
l/m
g p
rote
in)
MD-d8 MD-d8MD-d8MD-d8
40
50
60
70
80
90
Co
nce
ntr
ati
on
(
pm
ol/
mg
pro
tein
) MK4-d7
MK4-d7 epoxide
∗∗∗
∗∗∗
Conversion of PK-d7 and MK-4-d12 into MK-4-d7 in Sf9 cells transfected
with siControl or pcDNA3.3-UBIAD1(UBIAD1 expression vector)
EtOH MD-d8 PK-d7 MK-4-d12
0
10
20
30
40
Co
nce
ntr
ati
on
(
pm
ol/
mg
pro
tein
)
EtOH MD-d8 PK-d7 MK-4-d12
Sf9-Control Sf9-UBIAD1 expression vector
∗∗∗
Values are means and s.e.m. (n=6). Three asterisks, P<0.001 versus control-infected Sf9 cells with the same
compound treatment.
UB
IAD
1 a
cti
vit
y
(pm
ol/m
g p
rote
in/m
in-1
)
20
25
30
35
40
30
40
50
Sf9-control
Sf9-UBIAD1
UB
IAD
1a
cti
vit
y(p
mo
l/m
g p
rote
in/m
in-1
)
Substrate: K3-d8 (1 µµµµM)
Sf9-control
Sf9-UBIAD1
MK-4 biosynthetic activity of UBIAD1 in microsomes prepared from
UBIAD1 baculovirus-infected Sf9 cells with geranylgeranyl
pyrophosphate (GGPP) and MD
UB
IAD
1
(pm
ol/m
g p
rote
in/m
in
0
5
10
15
20
Substrate: MD-d8
0.01 0.1 1 (µµµµM)0
10
20
0 0.01 0.1 1 (µµµµM)
GGPP
UB
IAD
1(p
mo
l/m
g p
rote
in/m
in
UBIAD1-GFPTransmitted ER-Tracker Red Merge
50 µm
UBIAD1-GFPTransmittedBODIPY-TR
ceramide Merge
Subcellular localization of UBIAD1 in MG-63 cells
50 µm
UBIAD1-GFPTransmitted
MG-63 cells stably transfected with a UBIAD1–GFP expression vector were stained with ER-tracker Red or
BODIPY-TR ceramide (red) and were detected by GFP fluorescence (green). Merged images of GFP
fluorescence and by ER-marker or Golgi-marker fluorescence are shown at the right. The control construct
(mock-GFP) showed a diffuse fluorescence throughout the cytoplasm.
Nakagawa K. et al., Nature 2010; 468:117-121.
actin
)
1.2
1.4
1.6
1.8
1400
1600
1800
2000
MK-4
MK-4 epoxide
500
600
700
(pm
ol/g組織
)
MK-4-d7
MK-4-d7epoxide
UBIAD1 mRNA MK-4-d7 biosynthetic activity
Amounts of MK-4 and its epoxide
UBIAD1 mRNA expression, MK-4 biosynthetic activity, concentrations of MK-4 and its epoxide
in tissues of female mice
相対比
(UB
IAD
1/β
-actin
)
0.0
0.2
0.4
0.6
0.8
1.0
1.2
0
200
400
600
800
1000
1200
濃度
(pmol/g組織)
epoxide
0
100
200
300
400
K3-d
8からの生成量
(pm
ol/g
PK
MK-4
Prenylation
UBIAD1
Side-chain cleavage
GGPP
MK-4
R =
R=
O
O 3
3
OP
O
OH
OP
O
OH
HO
O
O
R
3
UBIAD1 is a novel biosynthetic enzyme for MK-4 that may
have both side-chain cleavage and prenylation activities
MK-4
K3
Side-chain cleavage
R= H3
Identification of UBIAD1 as a novel human menaquinone-4 biosynthetic enzyme
Kimie Nakagawa, Yoshihisa Hirota, Natsumi Sawada, Naohito Yuge, Masato Watanabe, Yuri Uchino, Naoko Okuda, Yuka Shimomura, Yoshitomo Suhara & Toshio Okano
Department of Hygienic Sciences, Kobe Pharmaceutical University,
Nature 2010; 468:117-121.
O
O
D
D
D
CD3
D
D
K1-d7 K3-d8
Vitamin K homologues
O
O
D
D
D
CD3
D3
Mevalonic acid
Acetyl-CoA
Mevalonate pathway
HMG-CoA
HMG-CoA reductase
Statin
Bisphosphonate
MK-4 biosynthesis in tissues is decreased by the
treatments with statins and bisphosphonates
MK-4-d7
O
O
D
D
D
CD3
D3
UBIAD1
Geranylgeranyl pyrophosphate(GGPP)
Geranyl pyrophosphate(GPP)
Farnesyl pyrophosphate(FPP)
FPP synthase
PP
3
POO-
O
O O-
O-
O
OO
O
PO-
O-
O
P :P
phylloquinone
geranylgeranyl pyrophosphate
MK-4 Vitamin K cycle
MK-4(quinone form)
MK-4(hydroquinone
prenylation
Mevalonate pathway
Possible interaction of MK-4 biosynthesis, vitamin K cycle and vitamin K action
MK-4 synthetic Enz.
(UBIAD1)
SXR/PXR signaling
Biological functions
phytol
PKA/PKC signaling
VKDP(active form)
cycle
VKDP(inactive form)
GGCX CO2, O2MK-4-(epoxide
form)
(hydroquinone from)cleavage
Generation of Ubiad1 knockout miceA
Targeting vector
3.5 kb 1.1 kb 6.7 kb
1.7 kb
: loxP
: FRT
Ubiad1exon2
Ubiad1exon1
mTOR
exon1Wild-type allele
neoR
Cre recombinase
neoRDTA
SalI ClaII
NheI
SpeI
NotISacII
KpnI
Ubiad1-neofloxed allele
ATG
15001000
500300
(bp)
M.W.
+/− +/+ −/−Posi500 bp 100 bpB
Targeted allele
F RPCR primer
(A) Schematic presentation of ubiad1 genome, targeting
vector and disrupted Ubiad1 genome. (B) PCR genotyping of
Ubiad1+/+, Ubiad1+/− and Ubiad1−/− embryos. PCR
genotyping of tail DNA of Ubiad1+/+, Ubiad1+/− and
Ubiad1−/− embryos. Lane 1, positive controls for Ubiad1+/−
allele. Lane 2, PCR bands of Ubiad1+/− embryos. Lane 3,
PCR bands of Ubiad1+/+ embryos. Lane 4, PCR bands of
Ubiad1−/− embryos.
Nakagawa K, Okano T, et al, PLOS ONE 2014; 9: 1-12, e104078
Morphological examination of Ubiad1+/+, Ubiad1+/− and Ubiad1−/− embryos and weanling mice (postnatal day 1) from pregnant Ubiad1+/− mice orally administered CoQ10
6 )))) Liver15
Brain
E
Day 1
DUbiad1+/+ Ubiad1+/− Ubiad1−/−Ubiad1+/+ Ubiad1+/− Ubiad1−/−
E15.5
HEstaining
A
B
0
2
4
Rela
tiv
e r
ati
o
(Ub
iad
1/G
ap
dh
)))) Liver
+/++/++/++/+ +/+/+/+/−−−− −−−−////−−−−0
5
10
Re
lati
ve
ra
tio
(Ub
iad
1/G
ap
dh
) Brain
N.D.N.D.
+/++/++/++/+ +/+/+/+/−−−− −−−−////−−−−
*
*
GAPDH
UBIAD1
F Ubiad1+/++/++/++/+ Ubiad1+/+/+/+/−−−− Ubiad1−−−−////−−−−
staining
C
UBIAD1staining
Ubiad1-deficient mouse embryos failed to survive beyond embryonic day 7.5 exhibiting small-sized body and gasturation arrest !!!
Flox/+ Cre/+
+/++/+
Cre/+
Flox+/+
+/+
Flox/+
Cre/+
-/+
Cre/+
Cre
Cre Cre
××××
Nestin-cre
Generation of a neural cell specific Ubiad1-/- mouse
flox/- flox/+cre/+
flox/+flox/-cre/+
Flox/+ Flox/+
+/+ Flox/Flox Flox/+ Flox/+
××××
Impairment of brain function
Co-Workers(Kobe Pharmaceutical University)
Kimie Nakagawa, Yoshihisa Hirota, Natsumi Sawada, Yuri Uchino, Masato Watanabe,
Syusaku Kimura, Naoto Yuge, Naoko Okuda, Yuka Shimomura, Makiko YamaneYuka Shimomura, Makiko Yamane
Dept. Hygienic SciencesKobe Pharmaceutical Univ.
(Shibaura Institute of Technology)Yoshitomo Suhara
Thank you for your attention!