A precursor form of vascular endothelial growth factor arises by initiation from an upstream in-frame CUG codon

Vascular endothelial growth factor (VEGF) is a mitogen in physiological and pathological angiogenesis. Understanding the expression of different VEGF isoforms might be important for distinguishing angiogenesis in tissue development, vascular remodelling and tumour formation. We examined its expression and noted the presence of the isoforms VEGF₁₂₁ and VEGF₁₆₅ (121 and 165 residues long respectively) in fetal heart, lung, ovary, spleen, placenta and ovarian tumours. Unexpectedly, a 47kDa species predominated in fetal intestine and muscle. The presumed initiation site in VEGF is an AUG codon (AUG¹⁰³⁹), 1039nt from its main transcriptional start site. AUG¹⁰³⁹ is preceded in the 5′ untranslated region by an in-frame CUG at nt 499 (CUG⁴⁹⁹), which could produce the 47kDa form with a 180-residue N-terminal extension. We therefore assessed whether CUG⁴⁹⁹ functions as an initiator. CUG⁴⁹⁹ initiation produced the 47kDa VEGF₁₆₅ precursor, which was processed at two sites to yield VEGF and three N-terminal fragments. When CTG⁴⁹⁹ was mutated to CGC, the precursor and N-terminal fragments were barely detectable. Although the precursor form was predominant in VEGF₁₆₅, both CUG⁴⁹⁹ and AUG¹⁰³⁹ forms were found in VEGF₁₂₁. VEGF precursor induced neither the proliferation of human umbilical vein endothelial cells nor the expression of angiopoietin 2, which can be induced by, and act with, VEGF to induce tumour angiogenesis. The precursor also adheres to the extracellular matrix (ECM), suggesting that it might be a storage form for generating active VEGF in the cell or ECM. Alternate CUG⁴⁹⁹ and AUG¹⁰³⁹ initiation and processing of the inactive precursor and its products might be important in regulating angiogenesis.