Andres Dekanty, Lazaro Centanin, Pablo Wappner.
Fundación Instituto Leloir, Buenos Aires, ARGENTINA.
The transcriptional response to hypoxia is a highly conserved
mechanism mediated by the heterodimeric (
α/
β) transcription factor HIF (hypoxia inducible factor). We have
previously defined a hypoxia-responsive
system homologous to HIF in Drosophila melanogaster, being
the proteins Similar (Sima) and Tango (Tgo) the functional
homologues of HIF-
α and
β subunits, respectively. Sima is regulated by oxygen through the
gene fatiga (a prolyl hydroxylase) that behaves as an oxygen
sensor. To gain insights on the regulation of the transcriptional
response to hypoxia, a HIF-Responsive-Element
(HRE) luciferase reporter was stably transfected in Drosophila
S2 cells. Expression of the reporter is strongly induced in hypoxia
or upon exposure to Deferoxamine (DFO), in a Sima/Tgo-dependent
manner. The HRE-reporter was then used in a
genome wide dsRNA-based screen. The screen led
to the identification of novel genes presumably required for the
transcriptional response to hypoxia, among which we identified
components of the insulin-signaling
pathway. We have demonstrated both in cell culture and in living
embryos, that insulin is a potent activator of Sima-dependent
transcription. This effect depends on PI3K and TOR pathways and
involves accumulation of Sima protein as well as an increase of its
nuclear localisation. It has been shown that PI3K and TOR pathways
play a fundamental role in growth regulation. Increased activity
promotes growth, while diminished signaling leads to cell and body
size reduction. Interestingly, we found that fatiga
loss-of-function led to body size
reduction but fatiga sima double mutants display normal size.
Consistent with this, Sima flip-out
over-expression led to cell size
reduction, strongly suggesting that Sima is a cell autonomous
negative regulator of growth. |