RMI1 deficiency in mice protects from diet and
genetic-induced obesity
Akira Suwa
1
, Masayasu Yoshino
2
, Chihiro Yamazaki
3
, Masanori Naitou
2
, Rie Fujikawa
3
,
Shun-ichiro Matsumoto
2
, Takeshi Kurama
1
, Teruhiko Shimokawa
1
and Ichiro Aramori
2
1 Pharmacology Research Labs, Astellas Pharma Inc., Tsukuba, Ibaraki, Japan
2 Molecular Medicine Labs, Astellas Pharma Inc., Tsukuba, Ibaraki, Japan
3 Trans Genic Inc., Chuo-ku, Tokyo, Japan
Introduction
Obesity is a complex disorder and a major risk factor
for metabolic diseases such as type 2 diabetes mellitus,
hypertension and cardiovascular disease. This energy
balance disorder is controlled by multiple pathways.
Several genes are known to be responsible for obesity:
the genes obese (ob) [1], fat (fa) [2], agouti (ay) [3],

established using the exchangeable gene trap method, and examined the
effects of deficiency of the target gene on diet and genetic-induced obesity.
The mutant strain 0283, which has an insertion at the recQ-mediated gen-
ome instability 1 (RMI1) locus, possesses a number of striking features that
allow it to resist metabolic abnormalities. Reduced RMI1 expression, lower
fasting-blood glucose and a reduced body weight (normal diet) were
observed in the mutant mice. When fed a high-fat diet, the mutant mice
were resistant to obesity, and also showed improved glucose intolerance
and reduced abdominal fat tissue mass and food intake. In addition, the
mutants were also resistant to obesity induced by the lethal yellow agouti
(A
y
) gene. Endogenous RMI1 genes were found to be up-regulated in the
liver and adipose tissue of KK-A
y
mice. RMI1 is a component of the
Bloom’s syndrome gene helicase complex that maintains genome integrity
and activates cell-cycle checkpoint machinery. Interestingly, diet-induced
expression of E2F8 mRNA, which is an important cell cycle-related mole-
cule, was suppressed in the mutant mice. These results suggest that the reg-
ulation of energy balance by RMI1 is attributable to the regulation of food
intake and E2F8 expression in adipose tissue. Taken together, these find-
ings demonstrate that RMI1 is a novel molecule that regulates energy
homeostasis.
Abbreviations
AUC, area under the curve; A
y
, lethal yellow agouti; BLM, Bloom syndrome; RMI1, recQ-mediated genome instability 1.
FEBS Journal 277 (2010) 677–686 ª 2009 The Authors Journal compilation ª 2009 FEBS 677
screened gene-trapped mice to identify novel energy

through Cre-mediated integration. We isolated 100
trap mouse strains in this study. One of these lines was
the 0283 mutant strain, which exhibits a remarkable
obesity-resistant phenotype. All homozygous embryos
died; therefore heterozygous mice (RMI1+ ⁄ )) were
used for this study (RMI1 was identified as the target
gene of this mutant strain as described below). Body
and organ weights as well as plasma parameters
(Tables S2 and S3) were measured, and learning, mem-
ory and behavioral tests (Table S4) as well as histo-
pathological analysis (Table S5) were performed for 8-
week-old RMI1+ ⁄ ) mice fed normal laboratory chow.
Although RMI1+ ⁄ ) mice had a phenotype almost
equivalent to that of the wild-type (RMI1+ ⁄ +), body
weight and fasting-plasma glucose were significantly
lower in RMI1 + ⁄ ) mice (Table 1).
Resistance to diet-induced obesity in RMI1+/)
mice
Wild-type (RMI1+ ⁄ +) and heterozygous (RMI1+ ⁄ ))
littermate mice were created via in vitro fertilization
using a single RMI1+ ⁄ ) male. At 4 weeks of age, the
individually housed littermates were fed either a normal
diet or one in which 60% of the calories were from fat
(high-fat diet). These mice were kept for 14 weeks, and
monitored for body weight changes and food intake.
Initially, the male RMI1+ ⁄ ) mice weighed less than
their male RMI1+ ⁄ + littermates, and those fed a nor-
mal diet consistently weighed less than their RMI1+ ⁄ +
littermates during the entire 14 weeks (Fig. 1A). The
rate of weight gain was equivalent for both genotypes

)
Triglycerides
(mgÆdL
)1
)
HDL cholesterol
(mgÆdL
)1
)
LDL cholesterol
(mgÆdL
)1
)
Male RMI1+ ⁄ + 20.6 ± 0.2 120.7 ± 7.6 1.15 ± 0.22 32.0 ± 3.1 33.2 ± 2.6 60.5 ± 2.2
RMI1+ ⁄ ) 18.8 ± 0.4** 86.7 ± 2.8** 1.61 ± 0.50 40.3 ± 2.6 39.0 ± 1.2 65.3 ± 3.2
Female RMI1+ ⁄ + 17.2 ± 0.3 112.7 ± 6.1 0.68 ± 0.14 28.2 ± 3.6 26.8 ± 1.5 61.2 ± 4.0
RMI1+ ⁄ ) 15.5 ± 0.1 *** 80.3 ± 3.0*** 0.85 ± 0.25 26.5 ± 2.5 34.0 ± 1.7* 59.3 ± 3.4
RMI1 deficiency prevents diet and genetic-induced obesity A. Suwa et al.
678 FEBS Journal 277 (2010) 677–686 ª 2009 The Authors Journal compilation ª 2009 FEBS
did not differ between the two feeding conditions
(Fig. 2A). The blood glucose and plasma insulin con-
centrations in the fasted or fed state did not differ sig-
nificantly between RMI1+ ⁄ ) and RMI1+ ⁄ + mice at
14 weeks (Table 2). However, an oral glucose tolerance
test showed that diet-induced glucose intolerance
improved significantly in RMI1+ ⁄ ) mice (Fig. 2C,D).
Insulin levels did not differ between RMI1+ ⁄ + and
RMI1+ ⁄ ) mice in the oral glucose tolerance test
(Fig. 2E,F).
Resistance to KK- and KK-A

y
⁄ a,
respectively).
Between 7 and 14 weeks of age, both RMI1+ ⁄ ) a ⁄ a
and RMI1+ ⁄ ) A
y
⁄ a mice experienced a consistent and
significant reduction of body weight compared to their
wild-type littermates (Fig. 3A). Hyperphagia induced
by the A
y
mutation was significantly less in RMI1+ ⁄ )
mice than RMI1+/+. However, KK-crossed
RMI1+ ⁄ ) mice did not show altered food intake, even
though their body weight was reduced (Fig. 3B). The
intra-abdominal fat found in KK-crossed mice was not
present in RMI1+ ⁄ ) mice. Similarly, the fat found in
the KK-A
y
-crossed RMI1+/) mice was a tendency
towards reduction compared to KK-Ay F1 mice
(Fig. 3D). Additionally, the enlargement of the liver
observed in KK- and KK-A
y
crossed mice was signifi-
cantly reduced in RMI1+ ⁄ ) mice (Fig. 3C).
The fasted blood glucose concentration in
RMI1+ ⁄ ) a ⁄ a and RMI1+ ⁄ ) A
y
⁄ a mice was signifi-

*
*
**
**
6
8
2
4
0
012345678910111213
Number of days fed diet
Body weight gain (%)
5.0
4.5
RMI1+/+ ND
RMI1+/
–
ND
3.5
4.0
3.0
***
*
2.0
2.5
Daily food intake (g)
0 1 2 3 4 5 6 7 8 9 1011121314
Number of days fed diet
45
40

Body weight (g)
**
** **
**
**
**
10
0 1 2 3 4 5 6 7 8 9 10 11 12 13 14
Number of days fed diet
5.0
4.5
RMI1+/+ HF
RMI1+/– HF
3.5
4.0
***
*
*
*
3.0
**
**
**
***
*
*
*
*
*
*

FEBS Journal 277 (2010) 677–686 ª 2009 The Authors Journal compilation ª 2009 FEBS 679
inserted into the first exon of the RMI1 gene (Gen-
bank accession number NM_028904). We attempted to
confirm that RMI1 is the target gene of this mutant
mice using quantitative PCR. The RMI1 mRNA level
in the skeletal muscle, fat, hypothalamus and liver of
RMI1+ ⁄ ) mice was approximately half that in
RMI1+ ⁄ + mice (Fig. 4A), which indicates that RMI1
is the responsible gene for this mutant mouse strain.
Next we compared the expression levels of RMI1 in
various tissues from normal mice. RMI1 mRNA was
expressed ubiquitously in most tissues (Fig. S1). To
clarify the association between RMI1 and the develop-
ment of obesity, we examined the RMI1 mRNA levels
in KK-A
y
mice. Five-week-old KK-A
y
mice did not
exhibit the obese phenotype. Therefore, we compared
the RMI1 mRNA levels of KK-A
y
mice before
(5 weeks) and after (15 weeks) obesity was observable.
Interestingly, the RMI1 mRNA level increased signifi-
cantly in the liver and intra-abdominal fat of the obese
phenotype mice; however, that in the skeletal muscle
did not increase, and that in the subcutaneous fat actu-
ally decreased (Fig. 4B). These results suggested that
the level of RMI1 expression in the fat and liver is

1.5
2
0.5
1
Plasma insulin (ng·mL
–1
)
0
0 0.2 0.4 0.6
Time (h)
350
400
450
250
300
RMI1+/– HF
RMI1+/– ND
RMI1+/+ ND
RMI1+/+ HF
*
150
200
***
50
100
Blood glucose (mg·dL
–1
)
0
0 0.5 1 1.5 2

0.40
Plasma insulin AUC
0–2 h (ng·mL
–1
)
0.00
+/+ +/–
+/+ +/–
ND
HF
AB
CD
EF
Fig. 2. RMI1 heterozygous (RMI1+ ⁄ )) mice
had less visceral adipose tissue and lower
glucose tolerance than wild-type (RMI1+/+)
on a high-fat diet. Male wild-type (RMI1+ ⁄ +)
and mutant (RMI1+ ⁄ )) mice (n = 6 per
group) were fed a normal diet (ND) or a
high-fat diet (HF) for 14 weeks. (A)
RMI1+ ⁄ ) mice do not differ from RMI1+ ⁄ +
mice in terms of liver weight. (B) The
amount of intra-abdominal fat was signifi-
cantly less in RMI1+ ⁄ ) than RMI1+/+ fed a
high-fat diet than RMI1+/+. (C) The blood
glucose concentration during the oral glu-
cose tolerance test was significantly lower
in RMI1+ ⁄ ) mice than RMI1+/+ at 1 and
2 h after glucose injection. (D) The
RMI1+ ⁄ ) mice fed a high-fat diet had a

Normal diet RMI1+ ⁄ + 27.8 ± 0.9 123 ± 5.3 102 ± 6 1.7 ± 0.2 0.61 ± 0.05
RMI1+ ⁄ ) 24.5 ± 0.9* 125 ± 5.7 88 ± 7 1.5 ± 0.1 0.62 ± 0.04
High-fat diet RMI1+ ⁄ + 38.8 ± 0.8 140 ± 2.9 121 ± 3 3.6 ± 0.8 0.76 ± 0.06
RMI1+ ⁄ ) 31.8 ± 1.3** 156 ± 8.2 124 ± 9 3.2 ± 0.9 0.56 ± 0.08
1.4
1.6
**
P = 0.10
1.0
1.2
0.6
0.8
0.0
0.2
0.4
Intra-abdominal fat (g)
+/–+/+ +/–+/+
a/a A
y
/a
450
RMI1+/+ a/a
RMI1+/– a/a
300
350
400
RMI1+/+ A
y
/a
RMI1+/– A

0.5
1.0
+/+ +/– +/+ +/–
a/a A
y
/a
7.0
7.5
RMI1+/+ a/a
RMI1+/– a/a
RMI1+/+ A
y
/a
5.5
6.0
6.5
RMI1+/– A
y
/a
**
**
*
4.5
5.0
*
3.0
3.5
4.0
Daily food intake (g)
9 10111213

20
25
Body weight (g)
7 8 9 1011121314
Age (weeks)
700
*** *
500
600
**
300
400
100
200
Blood glucose AUC
0–2 h (mg·dL
–1
)
0
+/+ +/– +/+ +/–
a/a A
y
/a
AB
CD
EF
Fig. 3. RMI1 heterozygous (RMI1+ ⁄ )) mice
were resistant to the obesity, hyperphagia
and improved glucose intolerance induced
by the A

y
F
1
mice. Values
are means ± SEM. Asterisks indicate signifi-
cant differences: *P < 0.05, **P < 0.01
versus RMI1+ ⁄ +.
A. Suwa et al. RMI1 deficiency prevents diet and genetic-induced obesity
FEBS Journal 277 (2010) 677–686 ª 2009 The Authors Journal compilation ª 2009 FEBS 681
E2F1, 4 or 5 mRNA in mice fed a high-fat diet. In
contrast, E2F8 mRNA was strongly induced by high-
fat feeding (7.1-fold increase over mice fed a normal
diet). Interestingly, the expression of E2F8 mRNA
induced in RMI1+ ⁄ ) mice was much less (60% sup-
pression) than that in RMI1+ ⁄ + mice. Recent reports
have indicated that the E2F family quantitatively regu-
lates adipose cells and thus plays an important role in
the development of obesity [21]. These results suggest
that E2F8 is associated with development of obesity
via cell-cycle regulation in the metabolic tissues, and,
in this study, regulation of E2F8 was found to be med-
iated by RMI1.
Given that RMI1-deficient mice have been found to
eat significantly less food under conditions of excessive
energy diets than under normal conditions, we com-
pared levels of RMI1 mRNA in the hypothalamus
between normal and high-fat feeding conditions. The
results showed that RMI1 mRNA levels were signifi-
cantly higher in the hypothalamus under high-fat feed-
ing conditions than under normal feeding (Fig. 4C).

a ⁄ a RMI1+ ⁄ + 37.2 ± 0.7 147 ± 1.7 126 ± 4 0.34 ± 0.03 0.58 ± 0.07 53 ± 27 1405 ± 189 268 ± 40 161 ± 15
RMI1+ ⁄ ) 31.8 ± 0.4*** 165 ± 2.7*** 113 ± 4* 0.34 ± 0.02 0.85 ± 0.1* 56 ± 11 1699 ± 103 257 ± 39 139 ± 13
Ay ⁄ a RMI1+ ⁄ + 51.0 ± 0.9 440 ± 11 175 ± 9 0.44 ± 0.03 0.32 ± 0.02 164 ± 17 1987 ± 241 460 ± 61 182 ± 21
RMI1+ ⁄ ) 44.3 ± 0.4*** 431 ± 9.6 150 ± 6* 0.37 ± 0.04 0.38 ± 0.03 191 ± 29 2545 ± 269 525 ± 39 153 ± 7
140
120
RMI1+/+
RMI1+/–
80
100
60
20
40
0
Relative expression of RMI1
Muscle Fat Hypo Liver
300
250
**
200
*
100
150
50
0
Muscle
515515515515
Age (week)
Relative expression of RMI1
Sub-fat Abd-fat Liver

high-fat diet (HF) conditions (n = 8 per
group). Fast, 16 h fasted. Values are
means ± SEM. Asterisks indicate significant
differences: *P < 0.05, **P < 0.01.
RMI1 deficiency prevents diet and genetic-induced obesity A. Suwa et al.
682 FEBS Journal 277 (2010) 677–686 ª 2009 The Authors Journal compilation ª 2009 FEBS
Agouti-related protein, pro-melanin-concentrating hor-
mone and CPT1c) in the hypothalamus of RMI1-defi-
cient mice. No changes were noted in the expression
levels of these factors (Table S1).
Discussion
Using a random mutagenesis approach based on the
exchangeable gene trap method, we identified RMI1 as
a novel regulator of energy homeostasis. The attributes
of RMI1 heterozygous mice, which exhibited a typical
lean phenotype, observed in this study are as follows:
first, RMI1-deficient mice were resistant to obesity
resulting from a high-fat diet or genetics. Second,
RMI1-deficient mice fed a high-fat diet gained less
abdominal fat. Third, the RMI1-deficient mice ate sig-
nificantly less food under the excess energy feeding
conditions. Fourth, impaired glucose tolerance induced
by high-fat diet or genetic obesity was improved in the
RMI1-deficient mice. In addition, levels of RMI1
expression were higher in the abdominal fat, liver and
hypothalamus of obese model mice than normal mice.
We could not find any abnormalities in the RMI1-
deficient mice under normal conditions, except the
reduced body weight and lower fasting glucose. Of
note is the fact that the deficient mice showed a rate of

Obesity develops as the result of an imbalance
between energy intake and expenditure. The reduction
of energy expenditure leads to an increase in fat mass,
ultimately resulting in obesity. The increase in cell
number (preadipocyte proliferation) and cell size (adi-
pocyte hypertrophy) is thought to be responsible for
the increase in the fat mass [14,15]. The cell cycle plays
an important role in preadipocyte proliferation, and is
regulated by several cell cycle-related proteins. RMI1
is known to be a cell cycle-related molecule with the
ability to activate the cell-cycle checkpoint machinery
[10], and siRNA depletion of RMI1 results in the
suppression of cell proliferation [13].
Sakai et al. have shown that a deficiency in the Skp2
gene, which encodes a cell cycle-related molecule,
results in resistance to obesity due to inhibition of
preadipocyte proliferation without causing adipocyte
hypertrophy [17]. This was found to be the case in
both the high-fat diet and Ay-induced obesity models.
Interestingly, the Skp2 knockout phenotype is very
similar to that of RMI1+ ⁄ ); however, Skp2 mRNA
levels were not altered in RMI1+ ⁄ ) mice. Fajas et al.
demonstrated that the E2F protein family also plays a
central role in preadipocyte proliferation, and that
E2F1-deficient mice are resistant to obesity induced by
a high-fat diet (due to the suppression of fat mass
accumulation) [21]. In this study, we found that the
high-fat diet upregulated E2F8 expression, but not that
of E2F1, E2F3 or E2F5. Interestingly, E2F8 upregula-
tion was suppressed in RMI1+ ⁄ ) mice. Although the

liferation [26,27]. Recently, Hagemann et al. reported
that E2F8 has a novel function as a guanine nucleotide
exchange factor for heterotrimeric G proteins [28].
Given the disparity of these reports, elucidation of
E2F8’s functions and contribution to the regulation of
cell proliferation will require further experiments.
Increased energy intake also leads to an increase in
the fat mass, which ultimately results in obesity. The
deficiency in RMI1 significantly decreased the food
intake only under conditions of excessive energy diet.
These results suggest that regulation of the energy bal-
ance by RMI1 is due to changes in the food intake.
Peripheral secreted adipocytokines, such as leptin, can
regulate food intake via the central nervous system in
response to changes in body fat content [29]. It is well
established that hypothalamic neurocircuits and signal
transductions modulate feeding behavior, thereby regu-
lating energy homeostasis [30]. First, we investigated
the expression levels of RMI1 in the hypothalamus.
The results showed that RMI1 expression was signifi-
cantly increased in the hypothalamus under high-fat
feeding conditions, and decreased under fasting condi-
tions. Next we examined whether these changes in
feeding behavior were based on modulation of central
nervous system pathways. Previous studies have shown
that several hypothalamic signaling factors, such as
neuropeptide Y and pro-opiomelanocortin, affect feed-
ing behavior via central nervous system pathways [30].
In the present study, we did not find any changes in
the expression levels of these factors; however, the pos-

y
experi-
ment, unfertilized eggs were collected from RMI1+ ⁄ )
females and fertilized in vitro with sperm from a KK-A
y
male (CLEA Japan). The genetic effects of the KK strain
and the A
y
mutation were investigated using F
1
mice. Only
male mice were used for the KK-A
y
experiment. For exami-
nation of the effects of high-fat feeding, 4-week-old mice
were fed a diet in which 60% of the calories were from fat.
The components of this high-fat diet were determined using
the method described by Ikemoto [32]. Briefly, the high-fat
diet contains 32% safflower oil, 33.1% casein, 17.6%
sucrose, 1.4% vitamin mixture, 9.8% mineral mixture, 5.6%
cellulose powder and 0.5% dl-methionine. Casein, sucrose
and the vitamin and mineral mixtures were purchased from
Oriental Yeast Co. Ltd (Tokyo, Japan), while the safflower
oil was purchased from Benibana Food (Tokyo, Japan) and
the dl-methionine from Wako Pure Chemical Industries
Ltd (Tokyo, Japan). The caloric density of this diet is
490 kcal per 100 g, with fat energy of 294.7 kcal per 100 g
(60.2%). The gonadal depots (representing intra-abdominal
fat) and liver tissues of of killed mice were removed and
weighed. All animal procedures were performed in accor-

for
the high-fat feeding study or 2.0 gÆkg
)1
for the KK-A
y
study), at time 0. Plasma samples for glucose measure-
ment were taken from the severed tail tips at 0.1, 0.5, 1.0
and 2 h.
Expression analysis of mRNA
The tissue samples were pulverized in liquid nitrogen, and
the total RNA was extracted using an Isogen kit (Nippon
Gene, Tokyo, Japan) according to the manufacturer’s
instructions. cDNAs were synthesized using SuperScript III
(Invitrogen, Carlsbad, CA, USA). Target mRNAs were
quantified via RT-PCR and the SYBR green method using
a PRISM 7900 sequence detector according to the manu-
facturer’s instructions (Perkin-Elmer Applied Biosystems,
Foster City, CA, USA). The level of mouse ribosomal pro-
tein (P0) was measured as an internal control. The primers
for each target gene are listed in Appendix S1.
Statistical analysis
The data represent the means ± SEM. The statistical sig-
nificance of the difference between groups was determined
using Student’s t test. P values < 0.05 were considered sig-
nificant. Statistical and data analyses were performed using
the sas 8.2 software package (SAS Institute Japan Ltd,
Tokyo, Japan).
Acknowledgements
We thank Drs Kiyoshi Furuichi, Masao Kato, Mas-
ayuki Shibasaki, Hitoshi Matsushime, Masato Kobori,

Terasaki H & Yamamura K (1999) Mutant mice lack-
ing Crk-II caused by the gene trap insertional mutagen-
esis: Crk-II is not essential for embryonic development.
Biochem Biophys Res Commun 266, 569–574.
8 Miyata K, Oike Y, Hoshii T, Maekawa H, Ogawa H,
Suda T, Araki K & Yamamura K (2005) Increase of
smooth muscle cell migration and of intimal hyperplasia
in mice lacking the a ⁄ b hydrolase domain containing 2
gene. Biochem Biophys Res Commun 329, 296–304.
9 Semba K, Araki K, Li Z, Matsumoto K, Suzuki M, Nak-
agata N, Takagi K, Takeya M, Yoshinobu K, Araki M
et al. (2006) A novel murine gene, Sickle tail, linked to the
Danforth’s short tail locus, is required for normal develop-
ment of the intervertebral disc. Genetics 172, 445–456.
10 Mankouri HW & Hickson ID (2007) The RecQ heli-
case–topoisomerase III–Rmi1 complex: a DNA struc-
ture-specific ‘dissolvasome’? Trends Biochem Sci 32,
538–546.
11 Raynard S, Zhao W, Bussen W, Lu L, Ding YY,
Busygina V, Meetei AR & Sung P (2008) Functional
role of BLAP75 in BLM-topoisomerase IIIa-dependent
Holliday junction processing. J Biol Chem 283, 15701–
15708.
12 Chang M, Bellaoui M, Zhang C, Desai R, Morozov P,
Delgado-Cruzata L, Rothstein R, Freyer GA, Boone C
& Brown GW (2005) RMI1 ⁄ NCE4, a suppressor of
genome instability, encodes a member of the RecQ heli-
case ⁄ Topo III complex. EMBO J 24, 2024–2033.
13 Yin J, Sobeck A, Xu C, Meetei AR, Hoatlin M, Li L &
Wang W (2005) BLAP75, an essential component of

218, 119–128.
21 Fajas L, Landsberg RL, Huss-Garcia Y, Sardet C, Lees
JA & Auwerx J (2002) E2Fs regulate adipocyte differen-
tiation. Dev Cell 3, 39–49.
22 Chester N, Kuo F, Kozak C, O’Hara CD & Leder P
(1998) Stage-specific apoptosis, developmental delay,
and embryonic lethality in mice homozygous for a tar-
geted disruption in the murine Bloom’s syndrome gene.
Genes Dev 12, 3382–3393.
23 Luo GSI, McDaniel LD, Nishijima I, Mills M, Yous-
soufian H, Vogel H, Schultz RA & Bradley A (2000)
Cancer predisposition caused by elevated mitotic recom-
bination in Bloom mice. Nat Genet 26, 424–429.
24 German J (1995) Bloom’s syndrome. Dermatol Clin 13,
7–18.
25 Wu L, Bachrati CZ, Ou J, Xu C, Yin J, Chang M,
Wang W, Li L, Brown GW & Hickson ID (2006)
BLAP75 ⁄ RMI1 promotes the BLM-dependent dissolu-
tion of homologous recombination intermediates. Proc
Natl Acad Sci USA 103, 4068–4073.
26 Christensen J, Cloos P, Toftegaard U, Klinkenberg D,
Bracken AP, Trinh E, Heeran M, Di Stefano L & Helin
K (2005) Characterization of E2F8, a novel E2F-like
cell-cycle regulated repressor of E2F-activated transcrip-
tion. Nucleic Acids Res 33 , 5458–5470.
27 Maiti B, Li J, de Bruin A, Gordon F, Timmers C,
Opavsky R, Patil K, Tuttle J, Cleghorn W & Leone G
(2005) Cloning and characterization of mouse E2F8,
a novel mammalian E2F family member capable of
blocking cellular proliferation. J Biol Chem 280, 18211–

Table S5. Histopathological findings.
Appendix S1. Primers for each target gene.
Appendix S2. Genomic DNA fragments obtained by
plasmid rescue.
This supplementary material can be found in the
online version of this article.
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RMI1 deficiency prevents diet and genetic-induced obesity A. Suwa et al.
686 FEBS Journal 277 (2010) 677–686 ª 2009 The Authors Journal compilation ª 2009 FEBS