Biology subjects

Korfhage, J. L.

Publications and source records attributed to Korfhage, J. L..

4 recordsLinked to original sources

bioRxiv2025

ATG2A engages Rab1a and ARFGAP1 positive membranes during autophagosome biogenesis

Autophagosomes form from seed membranes that expand through bulk-lipid transport via the bridge-like lipid transporter ATG2. The origins of the seed membranes and their relationship to the lipid transport machinery are poorly understood. Using proximity labeling and a variety of fluorescence microscopy techniques, we show that ATG2A localizes to extra-Golgi ARFGAP1 puncta during autophagosome biogenesis. ARFGAP1 itself is dispensable during macroautophagy, but among other proteins associating to these membranes, we find that RAB1 is essential. ATG2A co-immunoprecipitates strongly, albeit indirectly, with RAB1A, and siRNA-mediated depletion of RAB1A/B blocks autophagy downstream of LC3B lipidation, similar to ATG2A depletion. Further, when either autophagosome formation or the early secretory pathway is perturbed, ARFGAP1 and RAB1A accumulate at ectopic locations with autophagic machinery. Our results indicate that ATG2A engages a RAB1A complex on select early secretory membranes in support of autophagosome biogenesis. Significance StatementThis study expounds upon the role of early secretory membranes in autophagosome biogenesis. The authors demonstrate that RAB1/ARFGAP1-positive membranes are essential to autophagy and are recruited to the phagophore assembly site at an early step of autophagosome biogenesis. These membranes interact with the bridge-like lipid transport protein ATG2A and are positive for LC3B and WIPI2, suggesting that RAB1 membranes are a direct source for autophagosome formation.

cell biology↗

bioRxiv2023

Spartin-mediated lipid transfer facilitates lipid droplet turnover

Lipid droplets (LDs) are organelles critical for energy storage and membrane lipid homeostasis, whose number and size are carefully regulated in response to cellular conditions. The molecular mechanisms underlying lipid droplet biogenesis and degradation, however, are not well understood. The Troyer syndrome protein spartin (SPG20) supports LD delivery to autophagosomes for turnover via lipophagy. Here, we characterize spartin as a lipid transfer protein whose transfer ability is required for LD degradation. Spartin co-purifies with phospholipids and neutral lipids from cells and transfers phospholipids in vitro via its senescence domain. A senescence domain truncation that impairs lipid transfer in vitro also impairs LD turnover in cells while not affecting spartin association with either LDs or autophagosomes, supporting that spartins lipid transfer ability is physiologically relevant. Our data indicate a role for spartin-mediated lipid transfer in LD turnover. SignificanceThe Troyer syndrome protein spartin was proposed to function as a lipophagy receptor that delivers lipid droplets, organelles key for energy storage and membrane lipid homeostasis, to autophagosomes for degradation. We identify an additional function for spartin as a lipid transfer protein and show its transfer ability is required for lipid droplet degradation, including by lipophagy. Our data support that protein-mediated lipid transfer plays a role in lipid droplet turnover. Moreover, in spartins senescence domain we have discovered a new lipid transport module that likely also features in still undiscovered aspects of lipid droplet biology and membrane homeostasis.

cell biology↗

bioRxiv2023

ATG2A-mediated bridge-like lipid transport regulates lipid droplet accumulation

ATG2 proteins facilitate bulk lipid transport between membranes. ATG2 is an essential autophagy protein, but ATG2 also localizes to lipid droplets (LDs), and genetic depletion of ATG2 increases LD numbers while impairing fatty acid transport from LDs to mitochondria. How ATG2 supports LD homeostasis and whether lipid transport regulates this homeostasis remains unknown. Here we demonstrate that ATG2 is preferentially recruited to phospholipid monolayers such as those surrounding LDs rather than to phospholipid bilayers. In vitro, ATG2 can drive phospholipid transport from artificial LDs with rates that correlate with the binding affinities, such that phospholipids are moved much more efficiently when one of the ATG2-interacting structures is an artificial LD. ATG2 is thought to exhibit bridge-like" lipid transport, with lipids flowing across the protein between membranes. We mutated key amino acids within the bridge to form a transport-dead ATG2 mutant (TD-ATG2A) which we show specifically blocks bridge-like, but not shuttle-like, lipid transport in vitro. TD-ATG2A still localizes to LDs, but is unable to rescue LD accumulation in ATG2 knockout cells. Thus, ATG2 has a natural affinity for, and an enhanced activity upon LD surfaces and uses bridge-like lipid transport to support LD dynamics in cells.

cell biology↗

bioRxiv2021

Canagliflozin increases adenoma burden in female APCMin/+ mice

The diabetes drug canagliflozin acts primarily by inhibiting glucose reuptake by the sodium glucose transporter 2 (SGLT2) in the kidney proximal tubule, thereby lowering serum glucose levels. Canagliflozin also acts on SGLT1, a related transporter responsible for glucose uptake in the small intestine and more distal kidney tubules. Several cancers overexpress SGLT1 and SGLT2, where these transporters fuel tumor metabolism. A recent study by NIAs Interventions Testing Program (ITP) showed that canagliflozin treatment extends lifespan in male mice. Since cancer is the major cause of death in most mouse strains, including the UM-HET3 strain used by the ITP, this observation suggests that canagliflozin might exert anti-cancer effects in this context. Here, we treated a commonly-used mouse neoplasia model -- the intestinal adenoma-prone APCMin/+ strain -- with canagliflozin, to test the effects of drug treatment on tumor burden. Surprisingly, canagliflozin increased the total area of intestine involved by adenomas, an effect that was most marked in the distal intestine and in female mice. Immunohistochemical analysis suggested that canagliflozin may not influence adenoma growth via direct SGLT1/2 inhibition in neoplastic cells themselves. Instead, our results are most consistent with a model whereby canagliflozin aggravates adenoma development by altering the anatomic distribution of intestinal glucose absorption, as evidenced by increases in postprandial GLP-1 levels consistent with delayed glucose absorption. Our results suggest that canagliflozin exacerbates adenomatosis in the APCMin/+ model via complex, cell-non-autonomous mechanisms, and hint that sex differences in incretin responses may underlie differential effects of this drug on lifespan.

cancer biology↗