A  conserved ubiquitin- and ESCRT-dependent pathway internalizes human lysosomal membrane proteins for degradation

Abstract

The lysosome is an essential organelle to recycle cellular materials and maintain nutrient homeostasis, but the mechanism to down-regulate its membrane proteins is poorly understood. In this study, we performed a cycloheximide (CHX) chase assay to measure the half-lives of approximately 30 human lysosomal membrane proteins (LMPs) and identified RNF152 and LAPTM4A as short-lived membrane proteins. The degradation of both proteins is ubiquitin dependent. RNF152 is a transmembrane E3 ligase that ubiquitinates itself, whereas LAPTM4A uses its carboxyl-terminal PY motifs to recruit NEDD4-1 for ubiquitination. After ubiquitination, they are internalized into the lysosome lumen by the endosomal sorting complexes required for transport (ESCRT) machinery for degradation. Strikingly, when ectopically expressed in budding yeast, human RNF152 is still degraded by the vacuole (yeast lysosome) in an ESCRT-dependent manner. Thus, our study uncovered a conserved mechanism to down-regulate lysosome membrane proteins.

Safeguarding Lysosomal Homeostasis by DNAJC5/CSPα-Mediated Unconventional Protein Secretion and Endosomal Microautophagy

“…Endosomal microautophagy (eMI) refers to a special form of autophagy in which late endosomes or lysosomes take up cytoplasmic materials by membrane invagination and pinching off, forming multivesicular bodies (MVBs) (also called intralumenal vesicles) ( Marzella et al, 1981 ; Oku and Sakai, 2018 ). This process is conserved from S. cerevisiae to humans, involving several ESCRT machinery proteins ( Zhang et al, 2021 ). Because MVB formation is coupled to the engulfment of a portion of the cytosol into late endosomes, which is then degraded together with the invaginated membranes by lysosomes, eMI-mediated protein and membrane turnover appears to be largely non-selective.…”

GCAF(TMEM251) regulates lysosome biogenesis by activating the mannose-6-phosphate pathway

“…Our screen results highlighted the importance of the following functional groups: (1) M6P modification at the Golgi apparatus (GNPTAB, GNPTG, and MBTPS1), (2) endosomal trafficking, regulation, and fusion machinery (HOPS & CORVET components including VPS11/16/18/33A/39/41, LRRK2, and PIKFYVE), and (3) v-ATPase components on the lysosomes and its assembly factors (ATP6V0B/C/D1, ATP6V1A/B2/C1/D/G1, ATP6AP1/2, WDR7, and VMA21) (Fig. 1f ) 17 . Among the top 10 hits from the second round sorting, TMEM251, a gene of unknown function, stood out as the highest hit after RNF152, the reporter of the assay (Fig.…”

“…Stable cell lines were generated as described in Zhang et al 17 . In Brief, HEK293T cells were transfected with transfer plasmid, psPAX2 (Addgene 12260), and pMD2.G (Addgene 12259) at a 3.5:3.5:1 ratio using Lipofectamine 2000 according to the manufacturer’s instruction.…”

RNF152 negatively regulates Wnt/β-catenin signaling in Xenopus embryos

“…3 ), even though their respective mRNAs were injected at the same concentration. RNF152 has been shown to ubiquitinate itself, which leads to its short half-life ( 14 ). The steady-state protein level of E3 ligase-defective RNF152 was increased compared to that of WT RNF152 ( Fig.…”