Unconventional Trafficking of Mammalian Phospholipase D3 to Lysosomes

Abstract

Variants in the phospholipase D3 (PLD3) gene have genetically been linked to late-onset Alzheimer's disease. We present a detailed biochemical analysis of PLD3 and reveal its endogenous localization in endosomes and lysosomes. PLD3 reaches lysosomes as a type II transmembrane protein via a (for mammalian cells) uncommon intracellular biosynthetic route that depends on the ESCRT (endosomal sorting complex required for transport) machinery. PLD3 is sorted into intraluminal vesicles of multivesicular endosomes, and ESCRT-dependent sorting correlates with ubiquitination. In multivesicular endosomes, PLD3 is subjected to proteolytic cleavage, yielding a stable glycosylated luminal polypeptide and a rapidly degraded N-terminal membrane-bound fragment. This pathway closely resembles the delivery route of carboxypeptidase S to the yeast vacuole. Our experiments reveal a biosynthetic route of PLD3 involving proteolytic processing and ESCRT-dependent sorting for its delivery to lysosomes in mammalian cells.

PLD3 is a neuronal lysosomal phospholipase D associated with β-amyloid plaques and cognitive function in Alzheimer’s disease

“…Endolysosomal dysfunction in AD has become an important research focus, in part because dystrophic neurites are a consistent pathological feature of AD made up of enlarged axon segments around β-amyloid plaques clogged with dysfunctional lysosomes 24,25 . Here, we report PLD3 is highly concentrated in these abnormal neuronal lysosomes surrounding β-amyloid plaques (Figure 2), corroborating earlier reports of PLD3 upregulation around β-amyloid plaques 26 and that PLD3 can be routed to the lysosome 22 . The close and consistent association of PLD3 with β-amyloid neuropathology further strengthens the relevance of PLD3 to AD.…”

“…While most early reports suggested PLD3 was an endoplasmic reticulum-localized protein 21 , a few studies suggested it may, at least partially, localize to the lysosome. Most of these claims were based on studies of overexpression of the protein 22,23 . Because of contradictory reports regarding the subcellular localization of PLD3, we sought to better resolve the subcellular localization of PLD3 with attention to endogenous context-relevant tissue.…”

The lysosomal membrane—export of metabolites and beyond

“…Cleavage leads in this case to the release of the lumen-exposed, enzymatically active domains. LAP is synthesized as a type-I transmembrane protein [100], PLD3 as a type-II transmembrane protein [101]. Both proteins exhibit enzymatic activity (as acidic phosphatase and 5'-exonuclease, respectively) and are proteolytically cleaved in their luminal domains close to the transmembrane segments after arrival in lysosomes, releasing their larger enzymatically active domains as soluble enzymes.…”

“…In the case of LAP, the transmembrane form is transported from the trans-Golgi network (TGN) to the plasma membrane, from which it is internalized in endosomes before being finally delivered to dense lysosomes [100] (Figure 3B). PLD3 is sorted into intraluminal vesicles of multivesicular bodies after ubiquitination before being proteolytically cleaved and released as a soluble lysosomal enzyme [101]. In these examples, proteolytic processing is part of the biosynthetic delivery to lysosomes.…”

PLD3 epigenetic changes in the hippocampus of Alzheimer’s disease

“…PLD3 (phospholipase D family, member 3) ( OMIM * 615698) gene is located at chromosome 19q13.2 and encodes a lysosomal protein that belongs to the phospholipase D (PLD) superfamily, which catalyzes the hydrolysis of membrane phospholipids. However, PLD3 catalytic function has not yet been demonstrated [ 6 , 7 ]. PLD3 gene is highly expressed in the brain of healthy controls, particularly in several brain regions vulnerable to AD pathology, such as frontal, temporal, and occipital cortices and hippocampus, but reduced in neurons from AD brains [ 3 , 8 ].…”