Fig. 6

Accumulation of HTRA1 and its substrates in the CAA proteomic profile. a Immunofluorescence staining of an isolated capillary from a CAA patient (visualized by collagen IV staining) demonstrates a near complete co-localization of Aβ and HTRA1 immunoreactivity. b Venn diagram illustrating the overlap between proteins significantly altered in CAA with proteins significantly altered in CADASIL and in HTRA1 deficient mice as reported in our earlier study [72]. Eight of the shared proteins in CAA and CADASIL were also enriched in HTRA1 deficient mice (red filling). c Table with LFQ ratio information in the CAA, CADASIL and HTRA1 deficient proteomic profiles for the overlapping proteins, as for APCS, C3, C1QC and HTRA1. The latter four proteins are shared between CAA and CADASIL but not part of the HTRA1 deficient profile as they were not identified (APCS), not significantly altered (C3 and C1QC) or exclusively detected under HTRA1 wild-type conditions (HTRA1). Similar proteomic changes in HTRA1 deficient mice were recently reported by Kato et. al [34]. For most proteins in vitro cleavage data have been reported earlier. d, e APCS and PRSS23 are HTRA1 substrates. Shown are immunoblots of conditioned supernatants from HEK293 cells expressing PRSS23 (d) (detected via Myc-tag) or APCS (e), co-incubated with supernatants containing wild-type or active-site mutant (S328A) HTRA1 (detected via V5-tag) in the presence or absence of a HTRA1-specific inhibitor (5 μM). Molecular weight marker bands in kDa are indicated. The band above APCS in the S328A condition represent an unspecific cross-reactivity of the antibody. HTRA1 bands of lower molecular weight represent autoproteolysis fragments