Asialoglycoprotein receptor 2 (ASGR2, also known as hepatic lectin H2 or HL-2) is a 311-amino-acid single-pass type II transmembrane C-type lectin that serves as the minor subunit of the hepatic asialoglycoprotein receptor (Ashwell-Morell receptor) on the sinusoidal (basolateral) plasma membrane of hepatocytes. Sharing 58% sequence identity with the major subunit ASGR1 (H1), ASGR2 comprises an N-terminal cytoplasmic tail (residues 1-58) containing a phenylalanine-based internalization signal (FQDI, residues 5-8) and palmitoylated cysteine residues (Cys-54 and Cys-58), a transmembrane signal-anchor helix (residues 59-79), an extracellular coiled-coil stalk domain that mediates hetero-oligomerization with ASGR1, and a C-terminal extracellular C-type lectin carbohydrate-recognition domain (residues 176-302). Its carbohydrate-recognition domain carries a conserved QPD (Gln259-Pro260-Asp261) motif that recognizes terminal D-galactose and N-acetyl-D-galactosamine (GalNAc) units on desialylated plasma glycoproteins. Unlike ASGR1, which can assemble into functional homo-oligomers, ASGR2 requires co-expression and hetero-oligomerization with ASGR1 for efficient export from the endoplasmic reticulum to the plasma membrane, where the ASGR1/ASGR2 complex mediates clathrin-dependent receptor-mediated endocytosis of desialylated glycoprotein cargo for lysosomal degradation. When expressed in the absence of ASGR1 (particularly splice isoform H2a retaining the luminal juxtamembrane pentapeptide), unassembled ASGR2 precursors accumulate in the juxtanuclear endoplasmic reticulum quality control compartment and undergo Sec61-associated endoplasmic reticulum-associated degradation (ERAD).
| GO Term | Evidence | Action | Reason |
|---|---|---|---|
| GO:0004873 asialoglycoprotein receptor activity | TAS PMID:3863106 Sequence of a second human asialoglycoprotein receptor: cons... | ACCEPT | Summary: Traceable author statement from the cloning of human ASGR2 (H2) establishing its role as the second subunit of the hepatic asialoglycoprotein receptor. Reason: ASGR2 (H2) hetero-oligomerizes with ASGR1 (H1) to form the native hepatocyte asialoglycoprotein receptor complex and directly recognizes terminal D-galactose and N-acetyl-D-galactosamine (GalNAc) units on desialylated plasma glycoproteins via its QPD-motif C-type lectin domain. Supporting Evidence: PMID:3863106 From the same library, we have isolated and sequenced a clone encoding a second ASGP receptor, H2, with a protein sequence homology of 58% to H1. file:genes/human/ASGR2/ASGR2-uniprot.txt Mediates the endocytosis of plasma glycoproteins to which the terminal sialic acid residue on their complex carbohydrate moieties has been removed. The receptor recognizes terminal galactose and N-acetylgalactosamine units. |
| GO:0005515 protein binding | IPI PMID:11408579 A novel quality control compartment derived from the endopla... | REMOVE | Summary: Physical association of unassembled H2a precursor with ER translocon subunit Sec61b (Q9CQS8) during ERAD; bare GO:0005515 is removed per repository policy. Reason: While unassembled ASGR2 isoform H2a associates with the Sec61 translocon complex on its way to ER-associated degradation, bare GO:0005515 (protein binding) is uninformative and is removed under the repository's protein-binding policy. Supporting Evidence: PMID:11408579 On its way to degradation, H2a associated increasingly after synthesis with the ER translocon Sec61. |
| GO:0005515 protein binding | IPI PMID:11543633 Cloning, mapping, and characterization of a human homologue ... | REMOVE | Summary: Yeast two-hybrid and pull-down interaction with LASS2/CERS2 (Q96G23); bare GO:0005515 is removed per repository policy. Reason: Bare GO:0005515 (protein binding) from yeast two-hybrid and GST pull-down screening with LASS2/CERS2 (Q96G23) is uninformative and is removed under the repository's protein-binding policy. Supporting Evidence: PMID:11543633 Yeast two-hybrid screening and glutathione S-transferase pull-down assays showed that the LASS2 protein interacts with several membrane-associated receptors or transporters. |
| GO:0005515 protein binding | IPI PMID:21988832 Toward an understanding of the protein interaction network o... | REMOVE | Summary: High-throughput human liver yeast two-hybrid interaction with CERS2 (Q96G23); bare GO:0005515 is removed per repository policy. Reason: Bare GO:0005515 (protein binding) from high-throughput yeast two-hybrid interactome screening is uninformative and is removed under the repository's protein-binding policy. Supporting Evidence: PMID:21988832 In this study, we map the interactions of an unbiased selection of 5026 human liver expression proteins by yeast two-hybrid technology and establish a human liver protein interaction network (HLPN) composed of 3484 interactions among 2582 proteins. |
| GO:0005537 D-mannose binding | IBA GO_REF:0000033 | REMOVE | Summary: OVER_PROPAGATED: D-mannose binding propagated from PANTHER node PTN004568004 (seeded by CLEC17A/Q6ZS10); ASGR2 carries a galactose-specific QPD motif and specifically binds terminal D-galactose and N-acetyl-D-galactosamine rather than D-mannose. Reason: The PAINT annotation at PANTHER node PTN004568004 (within PTHR22803) is seeded by human CLEC17A (UniProtKB:Q6ZS10, prolectin), which binds terminal mannose and fucose. Unlike EPN-motif mannose-binding C-type lectins, ASGR2 contains the canonical galactose-recognizing QPD tripeptide (Gln259-Pro260-Asp261) in its carbohydrate-recognition domain and selectively recognizes terminal D-galactose and N-acetyl-D-galactosamine (GalNAc) on desialylated glycoproteins. Propagation Review Root cause: PROPAGATION BAD Failure modes: FUNCTIONAL DIVERGENCE WRONG ORTHOLOG OR PARALOG Sources checked: PANTHER:PTN004568004 · PTN004568004 SUPPORTS SOURCE BUT NOT TARGET Ancestral node in PANTHER family PTHR22803 combines mannose/fucose-binding and galactose/GalNAc-binding C-type lectin paralogs. UniProtKB:Q6ZS10 · CLEC17A SUPPORTS SOURCE BUT NOT TARGET Resolved in interpro/panther/PTHR22803/PTHR22803-entries.csv as human CLEC17A (prolectin), a mannose/fucose-binding C-type lectin whose glycan specificity does not transfer to the QPD-motif galactose/GalNAc receptor ASGR2. Supporting Evidence: file:genes/human/ASGR2/ASGR2-uniprot.txt Mediates the endocytosis of plasma glycoproteins to which the terminal sialic acid residue on their complex carbohydrate moieties has been removed. The receptor recognizes terminal galactose and N-acetylgalactosamine units. |
| GO:0005886 plasma membrane | IDA GO_REF:0000052 | ACCEPT | Summary: Immunofluorescence-based localization of ASGR2 to the plasma membrane. Reason: When co-expressed with ASGR1 in hepatocytes, ASGR2 incorporates into ASGR1/ASGR2 hetero-oligomers that are exported from the ER and localized to the sinusoidal plasma membrane. Supporting Evidence: PMID:29069408 This data parallels previous reports demonstrating that co-expression of ASGR1 results in ASGR2 incorporation into ASGR1/ASGR2 heterodimers that are then exported from the ER and are ultimately targeted to the plasma membrane |
| GO:0005886 plasma membrane | TAS Reactome:R-HSA-8855711 | ACCEPT | Summary: Reactome pathway assertion placing the palmitoylated ASGR1/ASGR2 receptor complex at the hepatocyte plasma membrane. Reason: Accurate core localization of the functional ASGR1/ASGR2 heteromeric receptor complex. Supporting Evidence: PMID:29069408 This data parallels previous reports demonstrating that co-expression of ASGR1 results in ASGR2 incorporation into ASGR1/ASGR2 heterodimers that are then exported from the ER and are ultimately targeted to the plasma membrane |
| GO:0006955 immune response | IBA GO_REF:0000033 | REMOVE | Summary: OVER_PROPAGATED: Immune response propagated from PANTHER node PTN001105054 seeded by myeloid C-type lectin pattern-recognition receptors (Clec4e, CLEC4D, Cd209b/d), FCER2, and invertebrate lectins; ASGR2 is a hepatocyte endocytic scavenger receptor subunit for endogenous desialylated plasma glycoproteins, not an immune receptor. Reason: PANTHER node PTN001105054 in family PTHR22803 groups hepatic asialoglycoprotein receptor subunits (ASGR1/ASGR2) with myeloid innate immune C-type lectin receptors (Clec4e/Mincle, CLEC4D/MCL, SIGN-R1/Cd209b, SIGN-R3/Cd209d), the low-affinity IgE Fc receptor FCER2 (P06734), and invertebrate/teleost C-type lectins. ASGR2 functions as the minor subunit of the hepatocyte Ashwell-Morell receptor that clears endogenous desialylated serum glycoproteins and aged desialylated platelets via clathrin-mediated endocytosis; this homeostatic glycoprotein and platelet turnover is not directly involved in an immune response (GO:0006955). Propagation Review Root cause: PROPAGATION BAD Failure modes: FUNCTIONAL DIVERGENCE WRONG ORTHOLOG OR PARALOG Sources checked: PANTHER:PTN001105054 · PTN001105054 SUPPORTS SOURCE BUT NOT TARGET Ancestral node in PANTHER family PTHR22803 spans both myeloid immune C-type lectins and hepatic asialoglycoprotein receptors. UniProtKB:P06734 · FCER2 SUPPORTS SOURCE BUT NOT TARGET Resolved in interpro/panther/PTHR22803/PTHR22803-entries.csv as human low-affinity IgE Fc receptor (FCER2/CD23), an immune receptor paralog whose immune function does not transfer to ASGR2. UniProtKB:Q8WXI8 · CLEC4D SUPPORTS SOURCE BUT NOT TARGET Resolved in interpro/panther/PTHR22803/PTHR22803-entries.csv as human CLEC4D (MCL), a myeloid innate-immune C-type lectin paralog. MGI:MGI:1298389 · Clec4e SUPPORTS SOURCE BUT NOT TARGET Corroborated in interpro/panther/PTHR22803/PTHR22803-entries.csv (Q9R0Q8, mouse Clec4e/Mincle); myeloid innate-immune C-type lectin paralog. MGI:MGI:1861231 · Cd209b SUPPORTS SOURCE BUT NOT TARGET Corroborated in interpro/panther/PTHR22803/PTHR22803-entries.csv (Q8CJ91, mouse Cd209b/SIGN-R1); innate-immune C-type lectin paralog. MGI:MGI:1861232 · Cd209d SUPPORTS SOURCE BUT NOT TARGET Corroborated in interpro/panther/PTHR22803/PTHR22803-entries.csv (Q91ZW8, mouse Cd209d/SIGN-R3); innate-immune C-type lectin paralog. Supporting Evidence: file:genes/human/ASGR2/ASGR2-uniprot.txt Mediates the endocytosis of plasma glycoproteins to which the terminal sialic acid residue on their complex carbohydrate moieties has been removed. The receptor recognizes terminal galactose and N-acetylgalactosamine units. |
| GO:0007166 cell surface receptor signaling pathway | TAS PMID:3863106 Sequence of a second human asialoglycoprotein receptor: cons... | REMOVE | Summary: Historical TAS annotation from the cDNA cloning paper PMID:3863106 conflating an endocytic cargo receptor with a cell-surface signaling receptor. Reason: PMID:3863106 reported the isolation and sequence of the human H2 (ASGR2) cDNA clone and did not assay or claim intracellular signal transduction. In GO, GO:0004873 (asialoglycoprotein receptor activity) is a transmembrane cargo receptor activity rather than a signaling receptor activity, and ASGR2 functions in clathrin-dependent receptor-mediated endocytosis of desialylated glycoproteins rather than a cell surface receptor signaling pathway. Supporting Evidence: PMID:3863106 From the same library, we have isolated and sequenced a clone encoding a second ASGP receptor, H2, with a protein sequence homology of 58% to H1. PMID:12171918 The hepatic asialoglycoprotein receptor (ASGP-R) internalizes desialylated glycoproteins via the clathrin-coated pit pathway and mediates their delivery to lysosomes for degradation. The human ASGP-R contains two subunits, H1 and H2. |
| GO:0009897 external side of plasma membrane | IBA GO_REF:0000033 | ACCEPT | Summary: Phylogenetic inference from PANTHER node PTN001105053 placing the receptor ectodomain on the external side of the plasma membrane. Reason: ASGR2 is a single-pass type II transmembrane protein whose C-terminal coiled-coil stalk and C-type lectin carbohydrate-recognition domain (residues 80-311) project into the extracellular space on the external side of the hepatocyte plasma membrane within the ASGR1/ASGR2 heteromeric complex. Propagation Review Root cause: NO FAILURE CORE Sources checked: PANTHER:PTN001105053 · PTN001105053 SUPPORTS TRANSFER Type II transmembrane topology with an extracellular C-terminal C-type lectin domain on the external face of the plasma membrane is conserved across this clade of cell-surface C-type lectin receptors. UniProtKB:Q9NNX6 · CD209 SUPPORTS TRANSFER Resolved in interpro/panther/PTHR22803/PTHR22803-entries.csv as human CD209 (DC-SIGN), a fellow type II cell-surface C-type lectin receptor. MGI:MGI:1916415 · Clec10a SUPPORTS TRANSFER Corroborated in interpro/panther/PTHR22803/PTHR22803-entries.csv (P49300, mouse Clec10a), a closely related galactose/GalNAc-specific type II cell-surface C-type lectin. MGI:MGI:95497 · Fcer2a SUPPORTS TRANSFER Corroborated in interpro/panther/PTHR22803/PTHR22803-entries.csv (P20693, mouse Fcer2), a fellow type II cell-surface C-type lectin. Supporting Evidence: file:genes/human/ASGR2/ASGR2-uniprot.txt Membrane; Single-pass type II membrane protein. PMID:29069408 This data parallels previous reports demonstrating that co-expression of ASGR1 results in ASGR2 incorporation into ASGR1/ASGR2 heterodimers that are then exported from the ER and are ultimately targeted to the plasma membrane |
| GO:0016020 membrane | IEA GO_REF:0000044 | MODIFY | Summary: Keyword-based mapping from UniProt Subcellular Location Membrane (SL-0162); refined to the more specific core localization GO:0005886 (plasma membrane). Reason: ASGR2 is a single-pass type II membrane protein that functions at the hepatocyte plasma membrane as part of the ASGR1/ASGR2 heteromeric receptor complex. Because GO:0016020 (membrane) is right but less specific than warranted by experimental evidence, it is refined to GO:0005886 (plasma membrane), which the gene already carries. Proposed replacements: plasma membrane Supporting Evidence: file:genes/human/ASGR2/ASGR2-uniprot.txt Membrane; Single-pass type II membrane protein. PMID:29069408 This data parallels previous reports demonstrating that co-expression of ASGR1 results in ASGR2 incorporation into ASGR1/ASGR2 heterodimers that are then exported from the ER and are ultimately targeted to the plasma membrane |
| GO:0016192 vesicle-mediated transport | IEA GO_REF:0000108 | MODIFY | Summary: Inter-ontology inference from GO:0004873 (asialoglycoprotein receptor activity) to broad GO:0016192 (vesicle-mediated transport); refined to the specific descendant GO:0006898 (receptor-mediated endocytosis). Reason: As the minor subunit of the hepatic asialoglycoprotein receptor complex, ASGR2 participates specifically in clathrin-dependent receptor-mediated endocytosis (GO:0006898) of desialylated plasma glycoproteins rather than generic vesicle-mediated transport. Proposed replacements: receptor-mediated endocytosis Supporting Evidence: PMID:12171918 The hepatic asialoglycoprotein receptor (ASGP-R) internalizes desialylated glycoproteins via the clathrin-coated pit pathway and mediates their delivery to lysosomes for degradation. The human ASGP-R contains two subunits, H1 and H2. file:genes/human/ASGR2/ASGR2-uniprot.txt Mediates the endocytosis of plasma glycoproteins to which the terminal sialic acid residue on their complex carbohydrate moieties has been removed. The receptor recognizes terminal galactose and N-acetylgalactosamine units. |
| GO:0038187 pattern recognition receptor activity | IBA GO_REF:0000033 | REMOVE | Summary: OVER_PROPAGATED: Pattern recognition receptor activity propagated from PANTHER node PTN001105054 seeded by myeloid innate-immune C-type lectin receptors; ASGR2 is a hepatocyte receptor subunit for endogenous desialylated plasma glycoproteins rather than a pathogen pattern-recognition receptor. Reason: PAINT propagated GO:0038187 (pattern recognition receptor activity) across PANTHER node PTN001105054 in family PTHR22803 from human and mouse myeloid pattern recognition receptors (CLEC6A/Dectin-2, CLEC4D/MCL, CLEC4E/Mincle, Cd209b/d). ASGR2 has functionally diverged as a hepatocyte-specific house-keeping receptor subunit (GO:0004873, asialoglycoprotein receptor activity) that recognizes terminal galactose/GalNAc on endogenous desialylated serum glycoproteins for clearance and does not function as a pattern recognition receptor. Propagation Review Root cause: PROPAGATION BAD Failure modes: FUNCTIONAL DIVERGENCE WRONG ORTHOLOG OR PARALOG Sources checked: PANTHER:PTN001105054 · PTN001105054 SUPPORTS SOURCE BUT NOT TARGET Ancestral node in PTHR22803 conflates myeloid PRR C-type lectins with hepatic asialoglycoprotein receptors. UniProtKB:Q6EIG7 · CLEC6A SUPPORTS SOURCE BUT NOT TARGET Resolved in interpro/panther/PTHR22803/PTHR22803-entries.csv as human CLEC6A (Dectin-2), a bona fide myeloid pattern-recognition receptor. UniProtKB:Q8WXI8 · CLEC4D SUPPORTS SOURCE BUT NOT TARGET Resolved in interpro/panther/PTHR22803/PTHR22803-entries.csv as human CLEC4D (MCL), a bona fide myeloid pattern-recognition receptor. UniProtKB:Q9ULY5 · CLEC4E SUPPORTS SOURCE BUT NOT TARGET Resolved in interpro/panther/PTHR22803/PTHR22803-entries.csv as human CLEC4E (Mincle), a bona fide myeloid pattern-recognition receptor. MGI:MGI:1861231 · Cd209b SUPPORTS SOURCE BUT NOT TARGET Corroborated in interpro/panther/PTHR22803/PTHR22803-entries.csv (Q8CJ91, mouse Cd209b/SIGN-R1), an innate-immune pattern-recognition receptor. MGI:MGI:1861232 · Cd209d SUPPORTS SOURCE BUT NOT TARGET Corroborated in interpro/panther/PTHR22803/PTHR22803-entries.csv (Q91ZW8, mouse Cd209d/SIGN-R3), an innate-immune pattern-recognition receptor. Supporting Evidence: file:genes/human/ASGR2/ASGR2-uniprot.txt Mediates the endocytosis of plasma glycoproteins to which the terminal sialic acid residue on their complex carbohydrate moieties has been removed. The receptor recognizes terminal galactose and N-acetylgalactosamine units. |
| GO:0042806 fucose binding | IBA GO_REF:0000033 | REMOVE | Summary: OVER_PROPAGATED: Fucose binding propagated from PANTHER node PTN004568004 (seeded by CLEC17A/Q6ZS10); ASGR2 carries a QPD galactose/GalNAc-recognition motif and does not bind fucose. Reason: PAINT propagated GO:0042806 (fucose binding) from human CLEC17A (UniProtKB:Q6ZS10) at PANTHER node PTN004568004. ASGR2 possesses the galactose-specific QPD motif (Gln259-Pro260-Asp261) in its C-type lectin carbohydrate-recognition domain and specifically recognizes terminal D-galactose and N-acetyl-D-galactosamine (GalNAc) rather than L-fucose. Propagation Review Root cause: PROPAGATION BAD Failure modes: FUNCTIONAL DIVERGENCE WRONG ORTHOLOG OR PARALOG Sources checked: PANTHER:PTN004568004 · PTN004568004 SUPPORTS SOURCE BUT NOT TARGET Ancestral node in PANTHER family PTHR22803 combines mannose/fucose-binding and galactose/GalNAc-binding C-type lectin paralogs. UniProtKB:Q6ZS10 · CLEC17A SUPPORTS SOURCE BUT NOT TARGET Resolved in interpro/panther/PTHR22803/PTHR22803-entries.csv as human CLEC17A (prolectin), whose fucose-binding specificity does not transfer to the QPD-motif galactose/GalNAc receptor ASGR2. Supporting Evidence: file:genes/human/ASGR2/ASGR2-uniprot.txt Mediates the endocytosis of plasma glycoproteins to which the terminal sialic acid residue on their complex carbohydrate moieties has been removed. The receptor recognizes terminal galactose and N-acetylgalactosamine units. |
| GO:0044322 endoplasmic reticulum quality control compartment | IDA PMID:11408579 A novel quality control compartment derived from the endopla... | KEEP AS NON CORE | Summary: Experimental localization of unassembled ASGR2 isoform H2a precursor to the juxtanuclear ER quality control compartment prior to ERAD when expressed without ASGR1. Reason: When expressed in the absence of its obligate hetero-oligomerization partner ASGR1, the precursor of ASGR2 isoform H2a accumulates in a specialized pre-Golgi endoplasmic reticulum quality control compartment (ERQC) en route to proteasomal degradation. Because this reflects quality-control retention of unassembled subunits rather than the mature receptor's site of function at the hepatocyte plasma membrane, it is retained as non-core. Supporting Evidence: PMID:11408579 We found that substrates of this pathway, the precursor of human asialoglycoprotein receptor H2a and free heavy chains of murine class I major histocompatibility complex (MHC), accumulate in a novel preGolgi compartment that is adjacent to but not overlapping with the centrosome, the Golgi complex, and the ER-to-Golgi intermediate compartment (ERGIC). |
| GO:0048471 perinuclear region of cytoplasm | IDA PMID:11408579 A novel quality control compartment derived from the endopla... | KEEP AS NON CORE | Summary: Juxtanuclear/perinuclear accumulation of unassembled ASGR2 isoform H2a in the ER quality control compartment in PMID:11408579; retained as non-core alongside GO:0044322. Reason: PMID:11408579 showed that unassembled H2a precursor, when expressed without its obligate assembly partner ASGR1 in heterologous cells, accumulates in a juxtanuclear/perinuclear pre-Golgi ER quality control compartment (also annotated to GO:0044322) prior to ERAD. Because GO:0044322 (endoplasmic reticulum quality control compartment) is not an ontology descendant of GO:0048471 (perinuclear region of cytoplasm) and this juxtanuclear accumulation reflects quality-control retention of unassembled precursor rather than the core hepatocyte cell-surface receptor localization, it is retained as KEEP_AS_NON_CORE. Supporting Evidence: PMID:11408579 We found that substrates of this pathway, the precursor of human asialoglycoprotein receptor H2a and free heavy chains of murine class I major histocompatibility complex (MHC), accumulate in a novel preGolgi compartment that is adjacent to but not overlapping with the centrosome, the Golgi complex, and the ER-to-Golgi intermediate compartment (ERGIC). |
| GO:0005534 galactose binding | TAS file:genes/human/ASGR2/ASGR2-uniprot.txt | NEW | Summary: Proposed NEW annotation capturing the direct terminal D-galactose and N-acetyl-D-galactosamine (GalNAc) binding specificity of the ASGR2 QPD-motif C-type lectin domain. Reason: Whereas GO:0005537 (D-mannose binding) and GO:0042806 (fucose binding) are over-propagated from EPN-motif C-type lectin paralogs in PANTHER family PTHR22803 and removed, ASGR2 carries a canonical galactose-recognizing QPD motif (Gln259-Pro260-Asp261) in its C-type lectin carbohydrate-recognition domain and specifically recognizes terminal D-galactose and N-acetyl-D-galactosamine (GalNAc) units on desialylated plasma glycoproteins. Supporting Evidence: file:genes/human/ASGR2/ASGR2-uniprot.txt Mediates the endocytosis of plasma glycoproteins to which the terminal sialic acid residue on their complex carbohydrate moieties has been removed. The receptor recognizes terminal galactose and N-acetylgalactosamine units. |
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Download this section (compressed HTML)Q: How does incorporation of ASGR2 into ASGR1/ASGR2 hetero-oligomers alter the spatial arrangement of C-type lectin carbohydrate-recognition domains and select for specific multi-antennary endogenous glycoprotein ligands in human hepatocytes?
Suggested experts: Martin Spiess, Paul H Weigel
Q: What physiological signals or hepatocyte states regulate alternative splicing of the ASGR2 luminal juxtamembrane pentapeptide (exon 3, VSP_003061) that governs ER quality control retention versus plasma membrane export?
Suggested experts: Gerardo Z Lederkremer, Martin Spiess
Experiment: Perform pulse-chase metabolic labeling, co-immunoprecipitation, and ER-to-Golgi transport assays across ASGR2 splice isoforms (P07307-1, P07307-2, P07307-3) in the presence and absence of ASGR1 in human hepatocyte lines.
Hypothesis: Hetero-oligomerization of ASGR2 with ASGR1 is required to mask an ER retention/ERAD degron in the juxtamembrane stalk region of ASGR2, enabling COPII-mediated ER export to the hepatocyte plasma membrane.
Type: Pulse-chase metabolic labeling and endo H sensitivity trafficking assay
Experiment: Compare clathrin-coated pit recruitment, internalization kinetics, and endosomal ligand dissociation in cells co-expressing wild-type ASGR1 with wild-type versus C54S/C58S mutant ASGR2.
Hypothesis: Dual S-palmitoylation of ASGR2 at Cys-54 and Cys-58 cooperates with Cys-36 palmitoylation on ASGR1 to drive partitioning of heteromeric ASGR1/ASGR2 complexes into clathrin-coated pits and pH-dependent uncoupling of ligands in sorting endosomes.
Type: Site-directed mutagenesis and quantitative endocytosis fluorescence microscopy
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