Hsp70 stabilizes lysosomes and reverts Niemann–Pick disease-associated lysosomal pathology

Abstract

Heat shock protein 70 (Hsp70) is an evolutionarily highly conserved molecular chaperone that promotes the survival of stressed cells by inhibiting lysosomal membrane permeabilization, a hallmark of stress-induced cell death. Clues to its molecular mechanism of action may lay in the recently reported stress- and cancer-associated translocation of a small portion of Hsp70 to the lysosomal compartment. Here we show that Hsp70 stabilizes lysosomes by binding to an endolysosomal anionic phospholipid bis(monoacylglycero)phosphate (BMP), an essential co-factor for lysosomal sphingomyelin metabolism. In acidic environments Hsp70 binds with high affinity and specificity to BMP, thereby facilitating the BMP binding and activity of acid sphingomyelinase (ASM). The inhibition of the Hsp70-BMP interaction by BMP antibodies or a point mutation in Hsp70 (Trp90Phe), as well as the pharmacological and genetic inhibition of ASM, effectively revert the Hsp70-mediated stabilization of lysosomes. Notably, the reduced ASM activity in cells from patients with Niemann-Pick disease (NPD) A and B-severe lysosomal storage disorders caused by mutations in the sphingomyelin phosphodiesterase 1 gene (SMPD1) encoding for ASM-is also associated with a marked decrease in lysosomal stability, and this phenotype can be effectively corrected by treatment with recombinant Hsp70. Taken together, these data open exciting possibilities for the development of new treatments for lysosomal storage disorders and cancer with compounds that enter the lysosomal lumen by the endocytic delivery pathway.

High sphingomyelin levels induce lysosomal damage and autophagy dysfunction in Niemann Pick disease type A

“…35 This possibility would be favored by lysosomal membrane permeabilization (LMP), to which NPA fibroblasts are prone under stress conditions. 36 In agreement, mild digitonin extraction allowing isolation of the cytosol from cells with intact organelles 37 indicated a 1.56-fold increase in the cytosolic and a concomitant 0.43-fold reduction in the cellular-organelle levels of the protease in the NPA fibroblasts compared with controls ( Figure 7c). To confirm that LMP results in accumulation of autophagolysosomes, we treated HeLa cells expressing EGFP-mRFP-LC3 with H 2 O 2 , which is a well-known inducer of lysosomal permeabilization.…”

“…Our findings showing LMP in NPA fibroblasts and neurons are in agreement with the beneficial effects shown for Hsp70 lysosomal delivery, which stabilized lysosomal membranes through ASM activation in NPA fibroblasts. 36 Our results indicate that direct modulation of SM levels could be an alternative/additional therapeutical strategy, which could be most relevant in patients in whom loss of function mutations preclude ASM activation. We find that the pharmacological activation of the neutral sphingomyelinase at the plasma membrane with dexamethasone prevents LC3-II accumulation as much as the general inhibition of sphingolipid synthesis with FB1.…”

Biochemical characterization of the interaction between HspA1A and phospholipids

“…This initial report was followed by many investigators showing that several members of the Hsp70 protein family, including HspA1A, as well as several bacterial Hsp70 (DnaK) proteins interact specifically with multiple glycolipids and phospholipids. These include phosphatidylserine (PS), bis-(monoacylglycero)-phosphate (BMP), globoyltriaosyl-ceramide (Gb3), sulfo-glycolipids, and anandamide (Arispe and De Maio 2000;Arispe et al 2002Arispe et al , 2004Armijo et al 2014;Broquet et al 2003;Browne et al 2007;Chen et al 2005;Gehrmann et al 2008;Harada et al 2007Harada et al , 2014Harada et al , 2015Kirkegaard et al 2010;Mahalka et al 2014;McCallister et al 2015;Oddi et al 2009;Petersen et al 2010;Whetstone and Lingwood 2003). These reports demonstrate that, although Hsp70s do not contain known lipid-binding domains, they interact with specific lipids and this interaction is evolutionarily conserved.…”

“…In addition to their indispensable functions in proteinchaperoning and cell signaling, Hsp70s also function at the plasma membrane (PM) and several organelle membranes, as well as in the extracellular environment (Broquet et al 2003;Gehrmann et al 2008;Kirkegaard et al 2010;Lancaster and Febbraio 2005a, b). Several members of the Hsp70 family, including HspA1A, the major heat-inducible Hsp70 in humans and mice, are translocated to the PM, are actively secreted from a variety of viable cells (Lancaster and Febbraio 2005a;Mambula et al 2007;Multhoff 2007), and bind to specific lipids as well as other surface molecules like acidic glycans (De Maio 2011, 2014Harada et al 2014).…”

Dynamics of endogenous Hsp70 synthesis in the brain of olfactory bulbectomized mice

“…It is also possible that Hsp70 induction may stimulate other systems that are responsible for cell survival after brain injury (Seidberg et al 2003;Robinson et al 2005). Cumulative indirect effects of Hsp70, such as suppression of apoptosis (Tidwell et al 2004), lysosome stabilization (Kirkegaard et al 2010), stimulation of the innate immune response (Johnson and Fleshner 2006;Gong et al 2009), suppression of the early preoligomeric stages of Aβ self-assembly (Evans et al 2006;Calderwood 2010), inhibition of proinflammatory signaling (Rozhkova et al 2010), and increase of survival of endogenous neural progenitor cells (Doeppner et al 2009) may account for the observed correlation. Possibly, Hsp70 induction somehow activates other compensatory mechanisms that facilitate regeneration and help to restore for some time normal functioning of neurons after the injury.…”