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Biology subjects

Kast, D. J.

Publications and source records attributed to Kast, D. J..

3 recordsLinked to original sources

MAPT regulates autophagic-lysosomal function and phagocytosis in human microglia

Tauopathies are characterized by the accumulation and spread of pathogenic tau aggregates throughout the brain, a process that is increasingly recognized to involve not only neurons but also microglia. However, whether pathogenic MAPT directly alters microglial degradative capacity remains poorly understood. Here, using isogenic human induced pluripotent stem cell-derived microglia carrying the pathogenic MAPT IVS10+16 mutation, we identify tau as a regulator of microglial lysosomal function. MAPT IVS10+16 microglia exhibited coordinated suppression of lysosomal and autophagic pathways, reduced lysosomal protease abundance and activity, and impaired autophagosome-lysosome fusion. Mutant microglia also showed reduced uptake of extracellular tau aggregates, reduced tau accumulation in acidic compartments, and a blunted lysosomal response to proteopathic stress. Conversely, genetic loss of MAPT increased lysosomal degradative capacity and accumulation of extracellular tau aggregates within acidic compartments, supporting a cell-intrinsic role for endogenous tau in regulating microglial degradative function. Pharmacologic enhancement of the autophagy lysosome pathway in MAPT IVS10+16 microglia increased proteolytic activity and improved tau handling. Together, these findings reveal a reciprocal relationship between tau and microglial lysosome function and identify degradative capacity as a modifiable component of the microglial response to tau pathology.

neuroscience↗

DFCP1 is a Regulator of ATGL-mediated Lipid Droplet Lipolysis

Lipid droplets (LDs) are transient lipid storage organelles that can be readily tapped to resupply cells with energy or lipid building blocks, and therefore play a central role in cellular metabolism. Double FYVE Domain Containing Protein 1 (DFCP1/ZFYV1) has emerged as a key regulator of LD metabolism, where the nucleotide-dependent accumulation of DFCP1 on LDs influences their size, number, and dynamics. Here we show that DFCP1 regulates lipid metabolism by directly modulating the activity of Adipose Triglyceride Lipase (ATGL/PNPLA2), the rate-limiting lipase driving the catabolism of LDs. We show through pharmacological inhibition of key enzymes associated with LD metabolism that DFCP1 specifically regulates lipolysis and, to a lesser extent, lipophagy. Consistent with this observation, DFCP1 interacts with and recruits ATGL to LDs in starved cells, irrespective of known regulatory factors of ATGL. We further establish that this interaction prevents dynamic disassociation of ATGL from LDs and thereby impedes the rate of LD lipolysis. Collectively, our findings indicate that DFCP1 primes ATGL on LDs to promote rapid LD catabolism.

cell biology↗

Mechanism of IRSp53 inhibition by 14-3-3

Filopodia are precursors of dendritic spines and polarized cell migration. The I-BAR-domain protein IRSp53 is an essential regulator of filopodia dynamics that couples Rho-GTPase signaling to cytoskeleton and membrane remodeling, playing essential roles in neuronal development and cell motility. Here, we describe a mechanism whereby phosphorylation-dependent inhibition of IRSp53 by 14-3-3 counters membrane binding and activation by Cdc42 or downstream cytoskeletal effectors. Phosphoproteomics, quantitative binding studies and crystal structures show that 14-3-3 binds to two pairs of phosphorylation sites in IRSp53. Using bicistronic expression we obtained a heterodimer of IRSp53 in which only one subunit is phosphorylated, and show that each subunit of the IRSp53 dimer independently binds a 14-3-3 dimer. A FRET-sensor assay developed using natively phosphorylated and 14-3-3-binding competent IRSp53 purified from mammalian cells reveals opposite conformational changes in IRSp53 upon binding of activatory (Cdc42, Eps8) vs. inhibitory (14-3-3) inputs.

biochemistry↗