MPPs and HtrA1 appear to interact directly.

Abstract

<p>(<b>A</b>) MPPs displayed differential activity in affecting The pattern of extracellular HtrA1 complexes (arrowheads) from HeLa cell low-serum conditioned medium is differentially altered by treatment with MPPs (25 mM, 37°C, 1 hr), as demonstrated when probed with polyclonal anti-HtrA1 antibody. (<b>B</b>) MPPs may induce conformational changes upon binding to HtrA1. The accessibility of N-terminal and C-terminal epitopes in the presence of MPPs was determined by ELISA and compared to DMSO controls. Error bars indicate the standard deviation. (<b>C</b>) Extracellular HtrA1 (arrows) could be precipitated from HEK-HtrA1 conditioned medium using HEMIN-conjugated agarose beads but not control, unconjugated agarose beads. (<b>D</b>) Competitive binding experiments using conditioned medium from HEK-HtrA1 or HeLa cells pre-incubated with MPPs revealed reduced recovery of HtrA1 (arrows) in the presence of competitor compounds when probed with polyclonal anti-HtrA1 antibody. (<b>E</b>) Degradation of Fibulin 5 in fixed HeLa cells treated with HtrA1 conditioned medium was enhanced in the presence of TPP, ZPP and PPP-IX. RMA: Rosmarinic acid, TPP: tin protoporphyrin IX, ZPP: zinc protoporphyrin IX, PPP-IX: protoporphyrin IX, HEM: HEMIN, CPP: cobalt protoporphyrin IX, CM: conditioned medium.</p