Stabilin-1 localizes to endosomes and the trans-Golgi network in human macrophages and interacts with GGA adaptors

Abstract

Stabilin-1 and stabilin-2 constitute a novel family of fasciclin domain-containing hyaluronan receptor homologues recently described by us. Whereas stabilin-1 is expressed in sinusoidal endothelial cells and in macrophages in vivo, stabilin-2 is absent from the latter. In the present study, we analyzed the subcellular distribution of stabilin-1 in primary human macrophages. Using flow cytometry, expression of stabilin-1 was demonstrated on the surface of interleukin-4/dexamethasone-stimulated macrophages (MPhi2). By immunofluorescence and confocal microscopy, we established that stabilin-1 is preferentially localized in early endosome antigen-1-positive early/sorting endosomes and in recycling endosomes identified by transferrin endocytosis. Association of stabilin-1 was infrequently seen with p62 lck ligand-positive late endosomes and with CD63-positive lysosomes but not in lysosome-associated membrane protein-1-positive lysosomes. Stabilin-1 was also found in the trans-Golgi network (TGN) but not in Golgi stack structures. Glutathione S-transferase pull-down assay revealed that the cytoplasmic tail of stabilin-1 but not stabilin-2 binds to recently discovered Golgi-localized, gamma-ear-containing, adenosine 5'-diphosphate-ribosylation factor-binding (GGA) adaptors GGA1, GGA2, and GGA3 long, mediating traffic between Golgi and endosomal/lysosomal compartments. Stabilin-1 did not bind to GGA3 short, which lacks a part of the Vps27p/Hrs/STAM domain. Deletion of DDSLL and LL amino acid motifs resulted in decreased binding of stabilin-1 with GGAs. A small portion of stabilin-1 colocalized with GGA2 and GGA3 in the TGN in MPhi2. Treatment with brefeldin A resulted in accumulation of stabilin-1 in the TGN. Our results suggest that stabilin-1 is involved in the GGA-mediated sorting processes at the interface of the biosynthetic and endosomal pathways; similarly to other GGA-interacting proteins, stabilin-1 may thus function in endocytic and secretory processes of human macrophages.

Microarray analysis of IFN-γ response genes in astrocytes

“…The other genes of interest involved in the vesicular pathways that were upregulated were VPS54, required for retrograde transport of Golgi membrane proteins from endosomal compartments and synapsin II which in involved in post-trans-Golgi trafficking pathways (Bustos et al, 2001, Maienschein et al, 1999Conibear and Stevens, 2000). Additionally the genes, stabilin-1, Rab6B, and the adaptor complex, AP-1, which are all involved in vesicular trafficking from the Golgi to the endosomes, were all down regulated (Kzhyshkowska et al, 2004;Opdam et al, 2000;Robinson, 2004). Thus although the effect on vesicular trafficking found in this study was modest, collectively these genes do suggest that in astrocytes IFN-γ may affect genes that are involved in regulation of membrane trafficking events in the vesicular trafficking pathway.…”

Identification of Biomarkers for Early Diagnosis of Acute Myocardial Infarction

“…STAB1 (signal transducer and activator of transduction 3), a multifunctional glycoprotein, was shown to be cardioprotective [Wang et al, 2009b]. It has been known that as a scavenger receptor, STAB1 recognizes acetylated lowdensity lipoproteins that can bind to macrophages playing a role in clearance and interacting with a variety of growth factors in angiogenesis [Adachi and Tsujimoto, 2002;Kzhyshkowska et al, 2004;Salmi et al, 2004]. The present research revealed that in blood samples at an early stage of AMI, STAB1 was significantly upregulated in healthy and stable coronary arteries at the first day of discharge; whereas it showed no difference in samples after 6 months of discharge.…”

Harnessing the Multifunctionality in Nature: A Bioactive Agent Release System with Self‐Antimicrobial and Immunomodulatory Properties

“…[ 56 ] Cells were fi xed for 10 min with 2% paraformaldehyde (PFA) in PBS, permeabilized for 15 min with 0.5% Triton X-100 in PBS, and followed by another 10 min fi xation with 4% PFA in PBS. Samples were blocked with 3% bovine serum albumin (BSA) in PBS for 1 h. Samples were incubated for 1 h and 30 min with primary antibodies followed by several 5 min washing steps with PBS.…”