Astrocyte Unfolded Protein Response Induces a Specific Reactivity State that Causes Non-Cell-Autonomous Neuronal Degeneration.

Recent interest in astrocyte activation states has raised the fundamental question of how these cells, normally essential for synapse and neuronal maintenance, become pathogenic. Here, we show that activation of the unfolded protein response (UPR), specifically phosphorylated protein kinase R-like endoplasmic reticulum (ER) kinase (PERK-P) signaling-a pathway that is widely dysregulated in neurodegenerative diseases-generates a distinct reactivity state in astrocytes that alters the astrocytic secretome, leading to loss of synaptogenic function in vitro. Further, we establish that the same PERK-P-dependent astrocyte reactivity state is harmful to neurons in vivo in mice with prion neurodegeneration. Critically, targeting this signaling exclusively in astrocytes during prion disease is alone sufficient to prevent neuronal loss and significantly prolongs survival. Thus, the astrocyte reactivity state resulting from UPR over-activation is a distinct pathogenic mechanism that can by itself be effectively targeted for neuroprotection.

Increased TIMP-3 expression alters the cellular secretome through dual inhibition of the metalloprotease ADAM10 and ligand-binding of the LRP-1 receptor.

The tissue inhibitor of metalloproteinases-3 (TIMP-3) is a major regulator of extracellular matrix turnover and protein shedding by inhibiting different classes of metalloproteinases, including disintegrin metalloproteinases (ADAMs). Tissue bioavailability of TIMP-3 is regulated by the endocytic receptor low-density-lipoprotein receptor-related protein-1 (LRP-1). TIMP-3 plays protective roles in disease. Thus, different approaches have been developed aiming to increase TIMP-3 bioavailability, yet overall effects of increased TIMP-3 in vivo have not been investigated. Herein, by using unbiased mass-spectrometry we demonstrate that TIMP-3-overexpression in HEK293 cells has a dual effect on shedding of transmembrane proteins and turnover of soluble proteins. Several membrane proteins showing reduced shedding are known as ADAM10 substrates, suggesting that exogenous TIMP-3 preferentially inhibits ADAM10 in HEK293 cells. Additionally identified shed membrane proteins may be novel ADAM10 substrate candidates. TIMP-3-overexpression also increased extracellular levels of several soluble proteins, including TIMP-1, MIF and SPARC. Levels of these proteins similarly increased upon LRP-1 inactivation, suggesting that TIMP-3 increases soluble protein levels by competing for their binding to LRP-1 and their subsequent internalization. In conclusion, our study reveals that increased levels of TIMP-3 induce substantial modifications in the cellular secretome and that TIMP-3-based therapies may potentially provoke undesired, dysregulated functions of ADAM10 and LRP-1.