Hypoxia-Inducible Factors and Adenosine Signaling in Vascular Growth - Extracellular ATP and Adenosine as Regulators of Endothelial Cell Function - page 112

uitination and subsequent degradation through the proteasomal pathway. Thus
siRNA-mediated knockdown of VHL or absence of functional VHL as in nat-
urally occurring mutations, can also prevent degradation of HIFs [13, 58]. In
addition, CoCl 2 mediated inhibition of pVHL binding to the hydroxylated HIFs
can also prevent degradation [74]. HIFs also can be hydroxylated by FIHs (Factor
Inhibiting HIFs) at specific asparagine residues (N803 of HIF-1 α and N847

of HIF-2 ) located in the C-terminal transactivation domain. Hydroxylation of

asparagine residues under non-hypoxic conditions inhibits transactivation func-
tion of HIFs by preventing interaction of HIFs with CBP/p300. Thus both
stabilization and functional activity of HIFs are maintained under hypoxic condi-
tions.
Hypoxia-inducible transcription factors HIF-1

α

bind the same con-
sensus DNA binding element and can upregulate a common set of genes involved
in cell growth, proliferation and angiogenesis. These include among others the vas-
cular endothelial growth factor (VEGF), its receptors including VEGFR-1 (Flt-1)
and VEGFR-2 (KDR or flk-1) (Fig. 7.1). While both HIF-1

α and HIF-2 α

α and HIF-2 α are impor-

tant in vascular growth and proliferation, HIF-2 α is perceived to have a greater role

because of its abundance in endothelial cells.

7.4 HIFs in Vascular Growth and Development

Endothelial cells are essential in embryonic vascular growth and development. A
number of receptors on endothelial cells regulate endothelial cell function. Mitogens
that act on these receptors are either secreted by endothelial cells themselves or
contributed by other cell types. Further, most of these receptors and their ligands
are regulated by the HIFs. Thus, altering the HIF pathway can profoundly affect
vascular growth in a number of models. For example, a HIF-1 knockout mouse

shows severely reduced vascular growth that results in embryonic lethality by E11
[37]. A HIF-2

α

null mouse, in 129 SV/ICR background, exhibited severe vascular
defects that died by E12.5 [60]. Similar observations were obtained in other HIF-2

α

α

knockout or knockdown models [14, 17, 50]. Interestingly, HIF-2 α null mouse in

some backgrounds did not show vascular defects [66, 72].
Several genes downstream of the HIFs can influence vascular growth. The most
common among these targets is the vascular endothelial growth factor (VEGF),
a known inducer of angiogenesis. Loss of even a single allele of VEGF, caused
abnormal vascular growth and embryonic lethality by E12 [9, 27]. Receptors
for VEGF are also regulated by HIFs in endothelial cells. VEGF has a higher
affinity for VEGFR-1 than for VEGFR-2. Disruption of VEGFR-1 (Flt-1) and
VEGFR-2 (flk-1 or KDR) in mice leads to embryonic lethality due to vascular
overgrowth in the former [28] and reduced vascular growth in the latter [67].
Therefore multiple parallel pathways operate to fine tune vascular growth and devel-
opment during embryonic life. Importantly, HIFs appear to be central regulators of
angiogenesis.