Hypoxia, a major inducer of angiogenesis, triggers major changes of gene expression at the transcriptional level. Furthermore, global protein synthesis is blocked while internal ribosome entry sites (IRES) allow specific mRNAs to be translated. Here we report the transcriptome and translatome signatures of (lymph)angiogenic genes in hypoxic HL-1 cardiomyocytes: most genes are not induced at the transcriptome-, but at the translatome level, including all IRES-containing mRNAs. Our data reveal activation of (lymph)angiogenic factor mRNA IRESs in early hypoxia. We identify vasohibin1 (VASH1) as an IRES trans-acting factor (ITAF) able to bind RNA and to activate FGF1 and VEGFD IRESs in hypoxia while it exhibits an inhibitory effect in normoxia. This suggests a generalized process of IRES-dependent induction of (lymph)angiogenic growth factors in early hypoxia, with a variable IRESome composition. IRES-dependent translation thus appears as a pivotal process to trigger new vessel formation in ischemic heart.
pTRIP-CRF1AL+ bicistronic lentivector with FGF1 IRES
The bicitronic lentivector plasmid codes for renilla luciferase LucR and firefly luciferase Luc+ under the control of the cytomegalovirus (CMV) promoter, with the human FGF1 mRNA IRES A (430 nt-long leader A, Martineau et al, Mol Cell Biol. 2004) between the two reporter genes. The LucR gene used here is a modified version of LucR where all the predicted splice donor sites have been mutated. pTRIP is a Delta-U3 plasmid allowing to produce self-inactivating (SIN) lentivectors.
pTRIP-CRF1AL+.ape
pTRIP-CRF1AL+.ape
pTRIP-CRFL+ bicistronic lentivector with FGF2 IRES
The bicitronic lentivector plasmid codes for renilla luciferase LucR and firefly luciferase Luc+ under the control of the cytomegalovirus (CMV) promoter, with the human FGF2 mRNA IRES (480 nt-long leader region, Vagner et al, Mol Cell Biol. 1995) between the two reporter genes. The LucR gene used here is a modified version of LucR where all the predicted splice donor sites have been mutated. pTRIP is a Delta-U3 plasmid allowing to produce self-inactivating (SIN) lentivectors.
pTRIP-CRFL+.ape
pTRIP-CRFL+.ape
pTRIP-CRVAaL+ bicistronic lentivector with VEGFA IRES a
The bicitronic lentivector plasmid codes for renilla luciferase LucR and firefly luciferase Luc+ under the control of the cytomegalovirus (CMV) promoter, with the human VEGFA mRNA 302 nt-long IRES a between the two reporter genes (Huez et al, Mol Cell Biol. 1998). The LucR gene used here is a modified version of LucR where all the predicted splice donor sites have been mutated. pTRIP is a Delta-U3 plasmid allowing to produce self-inactivating (SIN) lentivectors.
pTRIP-CRVAaL+.ape
pTRIP-CRVAaL+.ape
pTRIP-CRVAaL+.ape
pTRIP-CRVAbL+ bicistronic lentivector with VEGFA IRES b
The bicitronic lentivector plasmid codes for renilla luciferase LucR and firefly luciferase Luc+ under the control of the cytomegalovirus (CMV) promoter, with the human VEGFA 485 nt-long IRES b (Huez et al, Mol Cell Biol. 1998) between the two reporter genes. The LucR gene used here is a modified version of LucR where all the predicted splice donor sites have been mutated. pTRIP is a Delta-U3 plasmid allowing to produce self-inactivating (SIN) lentivectors.
pTRIP-CRVAbL+.ape
pTRIP-CRVAbL+.ape
pTRIP-CRhVCL+ bicistronic lentivector with VEGFC IRES
The bicitronic lentivector plasmid codes for renilla luciferase LucR and firefly luciferase Luc+ under the control of the cytomegalovirus (CMV) promoter, with the human VEGFC 419 nt-long IRES (Morfoisse et al, Cell Reports 2014) between the two reporter genes. The LucR gene used here is a modified version of LucR where all the predicted splice donor sites have been mutated. pTRIP is a Delta-U3 plasmid allowing to produce self-inactivating (SIN) lentivectors
pTRIP-CRhVCL+.ape
pTRIP-CRhVCL+.ape
pTRIP-CRhVDL+ bicistronic lentivector with VEGFD IRES
The bicitronic lentivector plasmid codes for renilla luciferase LucR and firefly luciferase Luc+ under the control of the cytomegalovirus (CMV) promoter, with the human VEGFD 507 nt-long IRES (Morfoisse et al, Cancer Res 2016) between the two reporter genes. The LucR gene used here is a modified version of LucR where all the predicted splice donor sites have been mutated. pTRIP is a Delta-U3 plasmid allowing to produce self-inactivating (SIN) lentivectors
pTRIP-CRhVDL+.ape
pTRIP-CRhVDL+.ape
pTRIP-CRMP2L+ bicistronic lentivector with c-myc IRES
The bicitronic lentivector plasmid codes for renilla luciferase LucR and firefly luciferase Luc+ under the control of the cytomegalovirus (CMV) promoter, with the human c-myc IRES (363 nt-long leader from P2 promoter, Nanbru et al, J. Biol. Chem.1997) between the two reporter genes. The LucR gene used here is a modified version of LucR where all the predicted splice donor sites have been mutated. pTRIP is a Delta-U3 plasmid allowing to produce self-inactivating (SIN) lentivectors
pTRIP-CRMP2L+.ape
pTRIP-CREL+ bicistronic lentivector with EMCV IRES
The bicitronic lentivector plasmid codes for renilla luciferase LucR and firefly luciferase Luc+ under the control of the cytomegalovirus (CMV) promoter, with the encephalomyocarditis virus (EMCV) 640 nt-long IRES (Elroy-Stein et al, PNAS 1989) between the two reporter genes. This IRES contains the T7 promoter in 5'. The LucR gene used here is a modified version of LucR where all the predicted splice donor sites have been mutated. pTRIP is a Delta-U3 plasmid allowing to produce self-inactivating (SIN) lentivectors
pTRIP-CREL+.ape
pTRIP-CRHL+ bicistronic lentivector with no IRES
The bicitronic lentivector plasmid codes for renilla luciferase LucR and firefly luciferase Luc+ under the control of the cytomegalovirus (CMV) promoter, with a hairpin between the two reporter genes (no IRES control). The LucR gene used here is a modified version of LucR where all the predicted splice donor sites have been mutated. pTRIP is a Delta-U3 plasmid allowing to produce self-inactivating (SIN) lentivectors
pTRIP-CRHL+.ape
LucR-mut
Renilla luciferase gene with silent point mutations on 5 AGGT splice donor sites. These mutations do not alter luciferase activity but prevent cryptic splicing events in bicistronic vectors, that could interfere with IRES activity measurement.
