Group leader: Sven Enerbäck
Members of the group: Zahra
Arani, Michal Bell,
Anna Cederberg, Mats Grände, Mikael
Nilsson, Sandra Rodrigo-Blomqvist, Gunilla
Petersson, Rickard Westergren and Hilmar
Our research efforts are directed towards the analysis of the developmental
programs in which winged helix genes contribute to correct pattern formation
and organogenesis. Towards achieving these goals, we use a technology
that specifically mutates any gene you choose in the mouse. This technology
employs the exchange of DNA sequences, by homologous recombination, between
exogenous, newly added DNA sequences and the cognate chromosomal DNA sequences
in embryo-derived mouse stem (ES) cells. This process is referred to as
"gene-targeting". The ES cells containing the desired targeting event
are then used to generate mouse germ line chimeras, thereby transferring
the alteration to subsequent mouse generations. We are now using this
technology to analyze the function of winged helix genes believed to mediate
important developmental decisions in the mouse. A similar approach is
also being employed to study problems of more immediate relevance to human
medicine. In particular, "gene-targeting" is being used to generate mouse
models for human genetic diseases. Such animals will permit a deeper analysis
of the pathogenesis of the genetic disease as well as provide appropriate
subjects for testing new therapeutic protocols including somatic gene
therapy. Eventually gene targeting should also become directly applicable
toward correcting genetic defects in humans by somatic gene therapy.
An example of a hypothetical
pathway regulated during development of the inner ear, currently under
investigation in our group:
Examples of ongoing
projects, grouped by gene names, has been entered
Enerbäck, MD, PhD
FoxI1 is a member of the forkhead family of winged helix transcriptional
regulators. Forkhead genes have previously been shown to be instrumental
during embryogenesis in mammals, in particular during development
of the nervous system. Mice with a targeted disruption of the FoxI1
locus exhibit circling behaviour, pathological swim test and abnormal
reaching response - all common findings in mice with vestibular dysfunction.
These animals also fail to elicit a Preyer reflex, in response to
a suprathreshold auditory stimulation, as seen in mice with profound
hearing impairment. Histological examination of the inner ear reveals
a gross structural malformation of the vestibular part of the inner
ear as well as the cochlea. These structures have been replaced by
a single irregular cavity in which neither proper semicircular ducts
nor cochlea can be identified. We also show that at day 9.5 post coitum
(p.c.) FoxI1 is exclusively expressed in the otic vesicle.
These findings, implicate FoxI1 as an early regulator necessary
for development of both cochlea and vestibulum and identify its human
homologue FoxI1 (also known as FREAC-6, Fkh10 or HFH-3) as
a previously unknown candidate deafness gene at 5q34.
derived from wild type mouse.
FoxI1-/- mice the entire cochlea is missing.
Rodrigo-Blomqvist , PhD student
type mouse E9.5 hybridized with a FoxI1 cRNA sense probe
in a whole-mount hybridization experiment. No hybridization
signal can be detected. The otic vesicle is marked with an arrow.
type mouse E9.5 hybridized with a Fkh10 cRNA antisense probe
in a whole-mount hybridization experiment. At this developmental
stage FoxI1 is exclusively expressed in the otic vesicle
Vidarsson , PhD student
FOXC2 is a winged helix gene that protects against obesity hypertriglyceridemia,
and insulin resistance Obesity, hyperlipidemia and insulin resistance
are common forerunners of non-insulin dependent diabetes mellitus
(NIDDM) a serious and increasingly prevalent disease in the industrialised
part of the world. Thrifty genes are thought to conserve energy
during periods of famine whereas they constitute a risk for developing
obesity related conditions, e.g. NIDDM, when energy is abundant.
The precise nature of such hypothetical thrifty genes is not known.
We have showed that the transcription factor gene FOXC2 is exclusively
expressed in white (WAT) and brown (BAT) adipose tissue during postnatal
life. In mice over-expressing this gene, solely in WAT and BAT,
intra-abdominal WAT has acquired a "brown fat-like" histology. The
intra-abdominal WAT depot is reduced (see below) and the interscapular
BAT is hypertrophic. Increased FOXC2 expression has a pleiotropic
effect on gene expression in BAT and WAT. For instance, the BAT
specific gene uncoupling protein-1 (ucp-1) is induced in intra-abdominal
WAT and there are also increased steady state levels of mRNAs encoding:
beta 1,2 and 3-adrenergic receptors, adipsin, CEBPalpha, PPARgamma,
PPARgamma coactivator-1 (PGC-1), insulin receptor, insulin receptor
substrate-1 (IRS-1), IRS-2, insulin responsive glucose transporter-4
(GLUT-4) and hormone sensitive lipase (HSL). As a consequence hereof,
total body lipid content has decreased from 30% to 10% (p<0.0004)
while no significant change in total body weight can be observed,
serum triglycerides are down by more than 50% (p<0.004), serum glucose
showed a significant 16% reduction (p<0.05), serum insulin levels
are decreased by approximately 50% (p<0.03) and levels of free fatty
acids (FFA) in serum are down by 27% (p<0.001). In an intravenous
glucose tolerance test, mice over-expressing FOXC2, display significantly
increased glucose elimination - a sign of increased insulin sensitivity.
Even though it is likely that other pathways/genes are involved,
the induction of ucp-1 in WAT, the hypertrophy of BAT, the decrease
of intra-abdominal WAT depots and the upregulation of mRNAs encoding
beta-adrenergic receptors suggest that an increased sensitivity
in a beta-adrenergic/cAMP/protein kinase A (PKA) pathway, at least
in part, can explain the findings in mice over-expressing FOXC2.
We have demonstrated a PKA isozyme switch with increased levels
of the regulative subunit RI-alpha. The PKA type I holoenzyme (RIalpha2C2)
binds cAMP with higher affinity and activates more easily than the
PKA type II enzyme, we have showed that such a shift in cAMP sensitivity
exists in adipocytes from animals with an increased expression of
FOXC2. Thus, the adipocytes will have a lower threshold for PKA
activation by adrenergic stimuli as compared with wild type littermates.
Since FOXC2, directly or indirectly, regulates triglyceride metabolism,
adrenergic sensitivity and insulin action in adipocytes, in the
way described here, it fulfils the criteria for an anti-thrifty
gene. To our knowledge FOXC2 is the only known gene that, in a concerted
action, can counteract most, if not all, of the symptoms associated
with obesity including hypertriglyceridemia and insulin resistance.
A likely consequence of such action would be prevention of NIDDM.
WAT depot is reduced in mice over-expressing
moreover the depot has acquired a "brown fat-like" histology.
We would like
to propose a model in which the degree of FOXC2 induction, in response
to a caloric load, perhaps sensed by cues like insulin and TNFalpha,
determines the efficiency by which calories are converted to triglycerides
(TGs). A strong induction would lead to a less efficient conversion,
with calories ending up as heat rather than stored as TGs
Bell, Post Doc
Mats Grände, Post Doc
Westergren, PhD student
Daniel Nilsson, PhD student
Zahra Arani, PhD student