Regulated Proteolytic Processing of Reelin through Interplay of Tissue Plasminogen Activator (tPA), ADAMTS-4, ADAMTS-5, and Their Modulators

Abstract

The extracellular signaling protein Reelin, indispensable for proper neuronal migration and cortical layering during development, is also expressed in the adult brain where it modulates synaptic functions. It has been shown that proteolytic processing of Reelin decreases its signaling activity and promotes Reelin aggregation in vitro, and that proteolytic processing is affected in various neurological disorders, including Alzheimer's disease (AD). However, neither the pathophysiological significance of dysregulated Reelin cleavage, nor the involved proteases and their modulators are known. Here we identified the serine protease tissue plasminogen activator (tPA) and two matrix metalloproteinases, ADAMTS-4 and ADAMTS-5, as Reelin cleaving enzymes. Moreover, we assessed the influence of several endogenous protease inhibitors, including tissue inhibitors of metalloproteinases (TIMPs), α-2-Macroglobulin, and multiple serpins, as well as matrix metalloproteinase 9 (MMP-9) on Reelin cleavage, and described their complex interplay in the regulation of this process. Finally, we could demonstrate that in the murine hippocampus, the expression levels and localization of Reelin proteases largely overlap with that of Reelin. While this pattern remained stable during normal aging, changes in their protein levels coincided with accelerated Reelin aggregation in a mouse model of AD.

Cleavage within Reelin Repeat 3 Regulates the Duration and Range of the Signaling Activity of Reelin Protein

“…2B), indicating that Pro-1244 is critically involved in the recognition motif of the unidentified N-t site protease secreted from cortical neurons. Although ADAMTS-4 is one of the candidate protease that catalyzes N-t cleavage (43,44), we found that ADAMTS-4 was able to weakly cleave Reelin-PD (Fig. 2C, lane 8).…”

“…We and another group reported recently that ADAMTS-4 can catalyze the N-t cleavage of Reelin (43,44). To determine whether Reelin-PD could be cleaved by ADAMTS-4, Reelin with a FLAG epitope between RR6 and RR7 (ReelinR6-F-R7, 43) and its PD version were used as substrates (Fig.…”

“…Abnormal N-t cleavage has been implicated in Alzheimer disease (41) and epilepsy (42). Recent studies from our laboratory and other laboratories indicate that a disintegrin and metalloproteinase with thrombospondin motifs (ADAMTS) 4 is able to cleave Reelin at the N-t site (43,44), although its contribution to Reelin metabolism in vivo remains uncertain.…”

Decisive role of Reelin signaling during early stages of Alzheimer’s disease

“…Interestingly, tissue plasminogen activator, shown to be induced by and involved in removal of Aβ aggregates (Tucker et al, 2000), cleaves Reelin at its C-terminal end (Krstic et al, 2012b) and by that modulate its binding to ApoER2 (Nakano et al, 2007). However, the biological significance of the proteolytic processing of Reelin might be more complex (Jossin et al, 2007, Chameau et al, 2009, and likely involves more downstream effects than only reducing binding of Reelin to its receptors, as previously proposed (Kohno et al, 2009).…”

Pathophysiological Function of ADAMTS Enzymes on Molecular Mechanism of Alzheimer’s Disease

“…Proteolysis of proteoglycans by ADAMTS enzymes can have destructive effects, producing matrices with disrupted cell-matrix communication, matrices with impaired fibrillary networks, or matrices with altered biomechanical properties [11]. After numerous comprehensive studies, ADAMTS expression has been found in the CNS [49][50][51][52][53][54][55][56][57][58][59][60] and is known to be changed in disease conditions [44,51,53].…”