KLK3/PSA and cathepsin D activate VEGF-C and VEGF-D

Abstract

Vascular endothelial growth factor-C (VEGF-C) acts primarily on endothelial cells, but also on non-vascular targets, for example in the CNS and immune system. Here we describe a novel, unique VEGF-C form in the human reproductive system produced via cleavage by kallikrein-related peptidase 3 (KLK3), aka prostate-specific antigen (PSA). KLK3 activated VEGF-C specifically and efficiently through cleavage at a novel N-terminal site. We detected VEGF-C in seminal plasma, and sperm liquefaction occurred concurrently with VEGF-C activation, which was enhanced by collagen and calcium binding EGF domains 1 (CCBE1). After plasmin and ADAMTS3, KLK3 is the third protease shown to activate VEGF-C. Since differently activated VEGF-Cs are characterized by successively shorter N-terminal helices, we created an even shorter hypothetical form, which showed preferential binding to VEGFR-3. Using mass spectrometric analysis of the isolated VEGF-C-cleaving activity from human saliva, we identified cathepsin D as a protease that can activate VEGF-C as well as VEGF-D.

eLife digest

The lymphatic system is composed of networks of vessels that drain fluids from the body’s tissues and filter it back into the blood. Growing these vessels depends on a factor known as VEGF-C, which is released in an inactive form and must be cut by enzymes before it can work. One enzyme that is known to activate the VEGF-C signal when the early embryo is developing is ADAMTS3. If this signal fails to switch on this can result in a condition known as lymphedema – whereby problems in the lymphatic system cause tissues to swell due to insufficient drainage. However, it is unknown whether the VEGF-C signal can be activated by enzymes other than ADAMTS3.

To investigate this Jha, Rauniyar et al. tested a specific family of proteins commonly found in the human prostate, which have previously been predicted to act on VEGF-C. This revealed that the lymphatic vessel growth factor can also be activated by an enzyme found in seminal fluid called prostate specific antigen, or PSA for short. To see if enzymes in other bodily fluids could switch on VEGF-C, different components of human saliva were separated and tested to see which could cut inactive VEGF-C. This showed that VEGF-C could be converted to an active form by another enzyme called cathepsin D.

Unexpectedly, Jha, Rauniyar et al. found that VEGF-C was also present in semen. For conception to occur PSA must liquify the semen following ejaculation. It was discovered that PSA activates VEGF-C just as the semen starts to liquify, suggesting that the lymphatic vessel growth factor might also play an important role in reproduction. In addition to VEGF-C, both PSA and cathepsin D were found to activate another growth factor called VEGF-D, which has an unknown role in the human body.

VEGF-C helps the spread of tumors, and blocking the two enzymes that activate this growth factor may be a new therapeutic approach for cancer. However, more work is needed to validate which types of tumor, if any, use these enzymes to activate VEGF-C. In addition, understanding the relationship between PSA and VEGF-C could help improve our knowledge of human reproduction.