Trafficking of a multifunctional protein by endosomal microautophagy: linking two independent unconventional secretory pathways

Abstract

During physiologic stresses, like micronutrient starvation, infection, and cancer, the cytosolic moonlighting protein glyceraldehyde‐3‐phosphate dehydrogenase (GAPDH) is trafficked to the plasma membrane (PM) and extracellular milieu (ECM). Our work demonstrates that GAPDH mobilized to the PM, and the ECM does not utilize the classic endoplasmic reticulum–Golgi route of secretion; instead, it is first selectively translocated into early and late endosomes from the cytosol via microautophagy. GAPDH recruited to this common entry point is subsequently delivered into multivesicular bodies, leading to its membrane trafficking through secretion via exosomes and secretory lysosomes. We present evidence that both pathways of GAPDH membrane trafficking are up‐regulated upon iron starvation, potentially by mobilization of intracellular calcium. These pathways also play a role in clearance of misfolded intracellular polypeptide aggregates. Our findings suggest that cells build in redundancy for vital cellular pathways to maintain micronutrient homeostasis and prevent buildup of toxic intracellular misfolded protein refuse.—Chauhan, A. S., Kumar, M., Chaudhary, S., Dhiman, A., Patidar, A., Jakhar, P., Jaswal, P., Sharma, K., Sheokand, N., Malhotra, H., Raje, C. I., Raje. M. Trafficking of a multifunctional protein by endosomal microautophagy: linking two independent unconventional secretory pathways. FASEB J. 33, 5626–5640 (2019). http://www.fasebj.org

Lysosome biology in autophagy

“…After being incorporated into intraluminal vesicles, eMI substrates can be degraded within endosomes or lysosomes 137 . They can even be secreted out of the cell 140 . A similar process has also been found in fission yeast 141 .…”

Autophagy in the Immunosuppressive Perivascular Microenvironment of Glioblastoma

“…Those forms of autophagy differ in how are regulated, the nature of the degraded substrates and in how the cargo is targeted to lysosomes. However, there is little data on the functions of endosomal microautophagy in tumor cells [24,25] or in the immune cells that could participate in the response to GB, macroautophagy and CMA are profusely reported to have important roles in those cell types. In this manuscript, we will review these two types of autophagy, focusing especially on CMA as an autophagy pathway that selectively degrades specific proteins that modulate tumor progression in the immunosuppressive perivascular microenvironment, which may constitute a promising target mechanism to treat this aggressive cancer.…”

Partial catalytic Cys oxidation of human GAPDH to Cys-sulfonic acid.

“…GAPDH is mainly a cytoplasmic enzyme, but it has been reported to moonlight as a cell surface and/or extracellular enzyme in healthy human cells 49,50 , in human cells subjected to stresses such as micronutrient starvation, hypoxia, infection, and cancer [50][51][52][53][54] , and to act as a extracellular effector in fungi and bacteria [55][56][57][58][59][60][61][62] . For recent reviews on GAPDH inhibitors and their multiple roles in several pharmacological applications see 63,64. In our study, HsGAPDH was purified from the supernatant of HEK293F cells treated with the mannosidase inhibitors kifunensine and kept for 4 days at 37°C after transfection with a DNA plasmid for recombinant expression of an endoplasmic reticulum glycoprotein.…”

“…GAPDH is mainly a cytoplasmic enzyme, but it has been reported to moonlight as a cell surface and/or extracellular enzyme in healthy human cells 49 , 50 , in human cells subjected to stresses such as micronutrient starvation, hypoxia, infection, and cancer 50 – 54 , and to act as a extracellular effector in fungi and bacteria 55 – 62 . For recent reviews on GAPDH inhibitors and their multiple roles in several pharmacological applications see 63 , 64 .…”