ARTICLE |
Correspondence to: Claire-Anne Gutekunst, Dept. of Neurology, Emory University, Woodruff Memorial Research Building, Suite 6000, 1639 Pierce Drive, Atlanta, GA 30322. E-mail: mguteku@emory.edu
![]() |
Summary |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
Stigmoid bodies (SBs) are structures in the cytoplasm of neurons. SBs are mostly found in the hypothalamic region of the rat and contain a protein called huntingtin-associated protein 1 (HAP1). In a recent publication, large cytoplasmic structures were shown to be immunoreactive for a type I receptor called SorLA/LR11. By light microscopic analysis, these structures appeared similar to SBs in size and in brain regional and subcellular localization. To determine whether these large puncta correspond to HAP1-containing SBs, we used antibodies specific to various domains of the apolipoprotein receptor LR11 to perform immunocytochemistry in rat and mouse brain tissue. Transfection studies using HeLa cells were conducted to demonstrate the specificity of the antibodies. We found that, in both species, antibodies to the domain II (or VSP10 for vacuolar sorting protein 10 domain) of LR11 immunoreact with large cytoplasmic structures. Co-localization immunolabeling experiments in rat brain tissue sections and in neuron cultures showed that these LR11-immunoreactive structures correspond to HAP1-positive SBs. Electron microscopy was performed in rat hypothalamus and further demonstrated the presence of LR11 in SBs and its co-localization with HAP1. LR11-containing SBs were most abundant in the hypothalamus but were also found in many brainstem nuclei, thalamus, and hippocampus. Our data also show that sortilin, another transmembrane protein containing a VPS10 domain, localizes to large cytoplasmic puncta and is found in LR11-positive and Hap1-positive SBs in hypothalamic neuron cultures. (J Histochem Cytochem 51:841852, 2003)
Key Words: huntingtin-associated protein 1, (HAP1), VPS10 domain, neurotensin receptor 3, nucleolus-like bodies
![]() |
Introduction |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
STIGMOID BODIES (SBs) are structures found in the cytoplasm of various types of neurons in the central and peripheral nervous systems (
In 1993, Shinoda and colleagues found that an antibody raised against a placental protein complex with aromatase activity (X-P2) detected SBs in the rat brain. X-P2-containing SBs were numerous in hypothalamic regions. However, the specific identity of the antigen present in the X-P2 complex that was detected in SBs was not determined. More recently, we have shown that multiple antibodies spanning the entire sequence of the huntingtin-associated protein 1 (HAP-1) detect SBs in rat brain (
A recent study determined that the apolipoprotein E receptor LR11 (lipoprotein receptor containing 11 LDL binding domains), also called SorLA (sorting protein-related receptor containing LDLR class A repeats), is localized to a large structure found in the perikarya of some neurons in the brains of rat and human (
![]() |
Materials and Methods |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
cDNA Construct and Transfection of Cells
PCR fragments of the rat HAP1A cDNA provided by Dr. Christopher Ross (nucleotides 51758 and 14211863 of GenBank
U38373) were cloned into PEGFP-C1 (ClonTech; Palo Alto, CA) by XhoI and KpnI. Full-length LR11 cDNA cloned into PCDNA3 was obtained from Dr. Chica Schaller. HeLa cells were used for transfection with LIPOFECTAMINE PLUS Reagent (Life Technologies, Lexington, KY). After 2448 hr, the cells were used for either immunocytochemistry (ICC) or Western blotting analysis.
Antibodies
A polyclonal antiserum specific for HAP1B (HAP1B-C, also called CAG1-3) was raised in rabbits against a synthetic peptide, EQQPIVPTQDSQRLE, corresponding to residues 595609 deduced from rat HAP1B cDNA sequence. Briefly, the peptide was coupled to keyhole limpet hemocyanin (Pierce; Rockford, IL) according to the manufacturer's instructions. Conjugated peptides were sent out to Pel Freeze (Redding, CA) where they were used as antigen to immunize rabbits and generate antisera (Pel Freeze). Antibodies were affinity-purified and characterized by ICC and Western blotting experiments (. The expressed fusion protein was purified and used for immunization of several female BALB/c mice. B-cells from draining lymph nodes were fused with PUA1 (ATCC; Rockville, MD) fusion partner and anti-fusion protein MAb-secreting cells were cloned by a dilution method. One of the clones, 1B6, showing specific immunoreactivity for both HAP1 isoforms by Western blots (
Western Blotting
Total homogenates from cultured cells and mouse brains were separated by SDS-PAGE, transferred to nitrocellulose membrane, and probed with the following antibodies: HAP1BC (1:1000), 1B6 (hybridoma medium), and mLR11 antibody (1:50). The proteins were detected by chemiluminescence (ECL system; Pierce). To determine that the lanes were loaded with equal amounts of proteins, blots were stained with Ponceau S before immunodetection.
Perfusion
Adult SpragueDawley rats (150200 g) and C57Bl6J mice (4050 g) were obtained from Jackson Laboratories (West Chester, PA) and maintained in a 12/12 light/dark cycle with ad libitum access to food and water. Rats (n=4) and mice (n=6) were deeply anesthetized with 4% choral hydrate solution (400 mg/kg). Perfusion was performed through the left cardiac ventricle with 34% paraformaldehyde (SigmaAldrich; St Louis, MO) in 0.1 M PB. For the electron microscopic analysis, rats (n=2) were perfused with 3% paraformaldehyde and 0.1% glutaraldehyde (Electron Microscopy Science; Fort Washington, PA). Brains were removed and postfixed overnight in 4% paraformaldehyde. Brains were cut in 10 series of 50-µm coronal sections using a vibratome. Sections were rinsed in 0.05 M PBS, pH 7.2, and processed for ICC as described below. All protocols involving animals were approved by the Emory University Animal and Care Committee and conform to NIH guidelines.
Neurospheres
Neurospheres were prepared by culturing cells from the hypothalamic region and colliculi and brainstem regions of Wistar rat fetuses. Fetuses of 14 days' gestation were obtained from pregnant rats under deep anesthesia with isoflurane. Tissue from the hypothalamus and brainstem were dissected under a dissecting microscope and dissociated into single cells by papain treatment, followed by gentle trituration with a fire-polished Pasteur pipette. The dissociated cells were cultured in suspension in 75-ml tissue culture flasks containing MEM with 10% fetal bovine serum, penicillin (100 U/ml; GIBCO, Grand Island, NY), streptomycin (100 g/ml; GIBCO). Within 35 days of culture at 37C with 5% CO2/95% air, cells proliferated and formed floating neurospheres. Neurospheres were then dissociated and plated on poly-lysine-coated coverslips and incubated in neurobasal medium supplemented with B27. In the differentiation medium, neurosphere cells attached to the dish, many of which extended a variety of cell processes.
Light Microscopic Single Immunolabeling
Free-floating sections were rinsed in PBS and processed for ICC. To reduce endogenous peroxidase activity and nonspecific antibody binding, sections were incubated in 3% hydrogen peroxide, rinsed several times in PBS, and incubated in PBS containing 4% normal goat serum (NGS) and 10 µg/ml avidin for 30 min. After PBS rinses, sections were incubated for 24 hr at 4C in 2% NGS in PBS containing 50 µg/ml biotin and either HAP1B-C (1:1000), mLR11 (1:200), anti-LR11 fibronectin domain (1:500), anti-LR11 C-terminal domain (1:1000), or rabbit anti-sortilin serum (SortK at 1:5000). Sections were then rinsed in PBS and incubated for 2 hr at 4C in biotinylated goat anti-mouse (mLR11) or goat anti-rabbit (all other antibodies) secondary antibodies used at 1:200 (Jackson Laboratories) in PBS containing 2% NGS. After rinses in PBS, sections then incubated in avidinbiotin complex (Vector ABC Elite; Vector Labs, Burlingame, CA) for 2 hr at 4C, followed by several PBS rinses. Final development was done by incubation in 0.05% in diaminobenzidine (DAB, Sigma) and 0.01% hydrogen peroxide in 50 mM Tris buffer for 515 min. Sections were rinsed thoroughly, mounted on glass slides, dehydrated, and coverslipped for light microscopic examination using a Leica DMRE. Brain localization was determined according to the rat brain atlas (
Double-label Immunofluorescence
Immunofluorescence was used to co-localize HAP1B and LR11 in rat brain and in primary neuron cultures and to co-localize sortilin with LR11 and HAP1 in primary cultures. Free-floating sections were rinsed in PBS, blocked in 5% NGS and 5% normal horse serum (NHS), and avidin for 60 min at room temperature (RT) and rinsed in PBS. After rinses in PBS, sections were incubated overnight at 4C in rabbit anti-HAP1B (HAP1B-C at 1:500) and mLR11 (1:50) in PBS containing 1% NHS, 1% NGS, and biotin. Sections were rinsed in PBS and incubated in biotinylated donkey anti-mouse IgG at 1:250 (Jackson Immunoresearch) in 1% NHS and 1% NGH for 1 hr at RT. Sections were rinsed in PBS and incubated in avidinbiotin complex (ABC kit; Vector Laboratories) for 60 min at RT. After rinses in PBS, sections were incubated in Cy5 tyramide (NEN; Boston, MA) according to the manufacturer's recommendation. After rinses, sections were incubated in FITC-conjugated donkey anti-rabbit (1:100) for 60 min at RT, rinsed in PBS, incubated for 30 min in 10 mM cupric sulfate in 50 mM ammonium acetate, pH 5.0, to eliminate autofluorescence, rinsed with PBS, then mounted on glass slides with Vectashield mounting medium for fluorescence. For some sections, nuclear counterstaining was obtained by a short incubation of the sections in PBS containing bis-benzimide before mounting. Staining of the primary cultures was done as follows. Coverslips were rinsed in cold PBS and fixed for 5 min in 4% paraformaldehyde. Coverslips were rinsed and blocked in 4% NGS for 30 min, rinsed, and incubated overnight at 4C in a combination of either HAP1BC(1:500) and mLR11 (1:50) antibodies, mLR11(1:50) and SortK (1:2500) antibodies, or 1B6(1:10) and SortK(1:2500). After several washes, coverslips were incubated with both rhodamine-conjugated goat anti-mouse and FITC-conjugated goat anti-rabbit secondary antibodies used at 1:100 (Jackson Immunoresearch). Nuclear counterstaining was obtained by a short incubation in PBS containing bis-benzimide before mounting. For each double-label experiment, controls included omission of one or both primary antibodies.
Electron Microscopy
For ultrastructural analysis, we used a combination of pre-embedded immunogold and DAB labeling of HAP1 and LR11, respectively. Single and double labeling were performed using HAP1BC and mLR11 antibodies. Sections were rinsed in PBS, blocked according to manufacturer's instructions, and incubated in HAP1B-C(1:1000), mLR11 (1:100), or a combination of HAP1B-C (1:1000) and mLR11 (1:100) overnight. Sections were rinsed in PBS and incubated in a combination of biotinylated donkey anti-rabbit and ultrasmall colloidal gold-conjugated secondary antibody (Aurion; Wageningen, The Netherlands) overnight. Sections were rinsed, incubated in avidinbiotin complex, and developed with DAB as described above. After a postfixation with 2.5% glutaraldehyde, gold particles in sections were intensified using an R-gent SE-EM silver enhancement kit (Aurion). Sections were then further fixed with 0.5% osmium tetroxide in 0.1 M PB for 15 min and processed for electron microscopy as described elsewhere (
![]() |
Results |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
HAP1 and Stigmoid Bodies
Our polyclonal antibody to HAP1BC identified a 110-kD protein in rat and mouse brain supernatant (Fig 1A), while our MAb to HAP1 (1B6) detected two distinct bands with molecular weights of approximately 110 kD and 95 kD (Fig 1A) using the rainbow molecular weight markers (BioRad; Hercules, CA). These molecular weights correspond to previously reported molecular weights for HAP1A and HAP1B respectively (
|
LR11 and Stigmoid Bodies
Most of the studies were performed using an LR11 MAb directed against the vacuolar protein sorting/targeting protein (VPS10) domain located in the extracellular region of LR11 (mLR11). mLR11 antibodies detect a unique protein band at a molecular weight of approximately 250 kD (Fig 1A) on Western blots of mouse brain homogenates. The characterization of this antibody is described elsewhere (
To further demonstrate the specificity of mLR11 antibodies and to rule out any crossreactivity between mLR11 and HAP1, we performed a series of experiments using transiently transfected HeLa cells. PEGFP-C1 containing either the HAP1 aa 1578 (EGFPHap1578), the HAP1A-specific C-terminal sequence (EGFPHAP1A-Ct), and PCDNA3 containing LR11 full-length cDNA (pSorLAOk) were transiently transfected into HeLa cells. After 24 hr, total homogenates from the transfected cells were blotted and immunoreacted for HAP1 or mLR11. HAP1 is mostly expressed by neurons, and HeLa cells do not show detectable levels of endogenous HAP1 expression. As expected, our 1B6 antibodies only detected EGFPHap1578 (Fig 2A) as a single band with a molecular weight of approximately 95 kD, which corresponds to the added weight of truncated HAP1A (60 kD) and GFP (
30 kD). LR11 MAbs detected endogenous and overexpressed full-length LR11 (Fig 2A) and did not react with EGFP-C1, EGFPHAP1578, or EGFPHAP1A-Ct. To verify that mLR11 did not crossreact with HAP1 proteins by ICC, HeLa cells were transiently transfected with a variety of HAP1- or LR11-containing vectors, fixed, and immunostained with 1B6 or mLR11. EGFP-transfected cells showed diffuse GFP in their cytoplasm and nuclei and reacted with neither 1B6 nor mLR11. EGFPHAP1578 induced the formation of 1B6-positive cytoplasmic inclusions that were negative for mLR11 (Fig 2B). The formation of the cytoplasmic inclusions after HAP1 transfection is consistent with our previous work in HEK 293 cells (
|
To determine whether the large cytoplasmic puncta immunodetected by the mLR11 antibodies did indeed correspond to the SBs detected by our various HAP1 antibodies, we performed double immunolabeling of rat brain tissue sections using HAP1BC and mLR11 antibodies. In the regions analyzed, including the lateral hypothalamus, the amygdala, the bed nucleus of the stria terminalis, and the cortical mantle, almost all of the HAP1BC-positive SBs were positive for mLR11 (Fig 3A). Double labeling was also performed at the electron microscopic level using mLR11 DAB staining and HAP1 immunogold. The ultrastructure of the mLR11 immunoreactive SBs was identical to that of HAP1-containing SBs. In double-stained sections, mLR11 DAB reaction product was present in HAP1 immunogold-positive SBs (Fig 3D).
|
SBs in the Rat and Mouse Brain
To further address the distribution of SBs in adult rat and mouse brain, we performed ICC using a variety of polyclonal and monoclonal HAP1 antibodies and monoclonal LR11 antibodies. Serial sections were cut in various planes (coronal, parasagittal, and horizontal) and immunoreacted. Both HAP1 and LR11 MAbs immunodetected SBs throughout the mouse and rat brain. Both markers detected very large numbers of SB-containing neurons in various nuclei of the hypothalamus, bed nucleus of the stria terminalis, amygdala, arcuate nucleus, subfornical organ, and the granule cell layer of the hippocampus. Regions in which both markers detected moderate numbers of SBs included nucleus accumbens, geniculate nucleus, septum, substantia nigra, and zona incerta. Regions with very few SBs included olfactory bulbs, striatum, globus pallidus, cortex, central gray, the habenula, and the cerebellum.
In our previous report of SBs using a fusion protein antibody against HAP1A and HAP1B, we had found little or no SBs in thalamic and hippocampal regions of the rat brain (
|
|
SBs in Neuron Cultures
To determine whether SBs also formed in primary neuron cultures, we performed HAP1 and LR11 ICC in primary neurosphere preparations from brainstem E14 rat embryos. Neurospheres were maintained in suspension for 5 days. They were then trypsinized and plated on poly-L-lysine-coated coverslips. Six days after plating, neurons were co-stained with polyclonal HAP1B and mLR11. HAP1/mLR11-co-immunoreactive SBs were visible as large puncta in the cytoplasm of neurons (Fig 6). These data represent the first description of SBs in cell cultures. The formation and presence of SBs in these cultures suggest that in vitro studies of SBs can be envisioned.
|
Sortilin in SBs
LR11 is a member of a recently identified type I receptor family that includes LR11, sortilin (also known as neurotensin receptor 3), SorCS1-3, and the yeast VPS10 protein (
![]() |
Discussion |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
In this study we set out to determine whether LR11 could be found in SBs. Here we present evidence showing that not only LR11 but also sortilin, another member of the VPS10 family of proteins, is present in SBs. Using well-characterized monoclonal and polyclonal antibodies generated against LR11 (
LR11 is a member of the LDL receptor gene family and has been shown to bind ApoE (
LR11, sortilin, and SorCS1 are type I membrane receptors with a small transmembrane spanning region mostly localized in intracellular vesicles and plasma membrane (
How can these transmembrane receptors reach SBs? What are the possible mechanisms leading to the presence of membrane proteins in SBs? Soluble forms of LR11 have been described in several studies. The ectodomain of mature LR11 can be shed from the transmembrane protein by a metalloprotease and has been detected in media from cell culture and brain slices using antibodies to the fibronectin domain but not to the intracellular domain (
Alternative routes that would take LR11 and sortilin to the cytosol and therefore make them accessible to SBs can be speculated. Both proteins could be in the cytoplasm as a result of misfolding and subsequent translocation from the endoplasmic reticulum (ER) to the cytosol. In recent years, several studies have shown the existence of a highly efficient quality control system in the ER, which allows the elimination of misfolded proteins or unassembled subunits of protein complexes. ER degradation of membrane proteins is a cytosolic event mediated by the proteasome (
Alternatively, the luminal region of SorLA/LR11 or sortilin could be cleaved once inside the endocytic compartment and thereafter translocated to the cytosol. Proteases including metalloproteinase, which have been shown to cleave SorLA, have been localized to early endosomes and could participate in such a mechanism (
In our previous studies we postulated that SBs were not aggregates but new subcellular organelles. HAP1A transfected into PC12 cells resulted in the formation of large inclusions (
SBs have been identified in neurons in various regions of the brain and have been detected by electron microscopy in the rodent brain as early as embryonic day 8 (
Whether the presence of HAP1, LR11, and sortilin in SBs indicates a role for these proteins in SB formation or in SB function remains unknown. We believe that identification of their molecular components will help shed light on their potential cellular function.
![]() |
Footnotes |
---|
1 Present address: Impfstoffwerk Dessau-Tornau GmbH, Viral Vaccines Production, Rosslau, Germany.
![]() |
Acknowledgments |
---|
Supported by a National Science Foundation grant IBN 9983078 to CAG.
We would like to thank Carlos Saavedra and Julie Jun for their technical assistance. We also thank Dr Christopher Ross for providing the rat HAP1A cDNA, Dr Chica Schaller for providing the PsorLAok plasmid, and Dr Kostya Kandror for providing the anti-sortilin serum.
Received for publication August 19, 2002; accepted December 20, 2002.
![]() |
Literature Cited |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
Bercovich Z, RosenbergHasson Y, Ciechanover A, Kahana C (1989) Degradation of ornithine decarboxylase in reticulocyte lysate is ATP-dependent but ubiquitin-independent. J Biol Chem 264:15949-15952
Blazquez J, Pastor F, Amat P, Pelaez B, Sanchez A, AmatPeral G (1995) Giant granular filamentous bodies in the cytoplasm of arcuate nucleus neurons of castrated rats. Histol and Histopathol 10:385-392
Bonifacino JS (1996) Reversal of fortune for nascent proteins. Nature 384:405-406[Medline]
Chabry J, Gaudriault G, Vincent JP, Mazella J (1993) Implication of various forms of neurotensin receptors in the mechanism of internalization of neurotensin in cerebral neurons. J Biol Chem 268:17138-17144
Chan E, Nassir J, Gutekunst CA, Coleman S, Maclean A, Mass A, Metzler M et al. (2002) Targeted disruption of huntingtin-associated protein-1 (Hap1) results in postnatal death due to depressed feeding behavior. Hum Mol Genet 11:945-959
Coux O, Tanaka K, Goldberg AL (1996) Structure and functions of the 20S and 26S proteasomes. Annu Rev Biochem 65:801-847[Medline]
Dennes A, Madsen P, Nielsen MS, Petersen CM, Pohlmann R (2002) The yeast Vps10p cytoplasmic tail mediates lysosomal sorting in mammalian cells and interacts with human GGAs. J Biol Chem 277:12288-12293
Fechner J (1986) Nucleolus-like bodies in the pineal gland of the Djungarian hamster (Phodopus sungorus). Cell Tissue Res 243:441-443[Medline]
Grillo M (1970) Cytoplasmic inclusions resembling nucleoli in sympathetic neurons of adults rats. J Cell Biol 45:100-117
Gutekunst C, Li S, Yi H, Ferrante R, Li X, Hersch S (1998) The cellular and subcellular localization of huntingtin-associated protein 1 (HAP1): comparison with huntingtin in rat and human. J Neurosci 18:7674-7686
Hampe W, Rezgaoui M, HermansBorgmeyer I, Schaller HC (2001) The genes for the human VPS10 domain-containing receptors are large and contain many small exons. Hum Genet 108:529-536[Medline]
Hampe W, Riedel IB, Lintzel J, Bader CO, Franke I, Schaller HC (2000) Ectodomain shedding, translocation and synthesis of SorLA are stimulated by its ligand head activator. J Cell Sci 113:4475-4485
Hermey G, Riedel IB, Rezgaoui M, Westergaard UB, Schaller C, HermansBorgmeyer I (2001) SorCS1, a member of the novel sorting receptor family, is localized in somata and dendrites of neurons throughout the murine brain. Neurosci Lett 313:83-87[Medline]
Hiller MM, Finger A, Schweiger M, Wolf DH (1996) ER degradation of a misfolded luminal protein by the cytosolic ubiquitin-proteasome pathway. Science 273:1725-1728
HindelangGertner C, Stoeckel M-E, Porte A, Dellmann H-D, Madarasz B (1974) Nematosomes or nucleolus-like bodies in hypothalamic neurons, the subfornical organ and adenohypophysial cells of the rat. Cell Tissue Res 155:211-219[Medline]
Jacobsen L, Madsen P, Nielsen MS, Geraerts WP, Gliemann J, Smit AB, Petersen CM (2002) The sorLA cytoplasmic domain interacts with GGA1 and -2 and defines minimum requirements for GGA binding. FEBS Lett 511:155-158[Medline]
Jiang A, Lehti K, Wang X, Weiss SJ, KeskiOja J, Pei D (2001) Regulation of membrane-type matrix metalloproteinase 1 activity by dynamin-mediated endocytosis. Proc Natl Acad Sci USA 98:13693-13698
Johnston JA, Johnson ES, Waller PR, Varshavsky A (1995) Methotrexate inhibits proteolysis of dihydrofolate reductase by the N- end rule pathway. J Biol Chem 270:8172-8178
Kandror KV, Pilch PF (1998) Multiple endosomal recycling pathways in rat adipose cells. Biochem J 331:829-835[Medline]
Katoh Y, Shimizu N (1982) Identity of Holmes positive bodies with electron microscopy demonstrate nucleolus-like bodies in neuronal cytoplasm. Stain Tech 57:83-89
Kopito RR (1997) ER quality control: the cytoplasmic connection. Cell 88:427-430[Medline]
LarrivaSahd J, Gorski R (1987) Ultrastructural characterization of the central component of the medial preoptic nucleus. Exp Neurol 98:370-387[Medline]
Leranth C, Shanabrough M, Naftolin F (1991) Estrogen induces ultrastructural changes in progesterone receptor-containing GABA neurons of the primate hypothalamus. Neuroendocrinology 54:571-579[Medline]
Li SH, Gutekunst CA, Hersch SM, Li XJ (1998a) Association of HAP1 isoforms with a unique cytoplasmic structure. J Neurochem 71:2178-2185[Medline]
Li SH, Gutekunst CA, Hersch SM, Li XJ (1998b) Interaction of huntingtin-associated protein with dynactin P150Glued. J Neurosci 18:1261-1269
Li SH, Li H, Torre ER, Li XJ (2000) Expression of huntingtin-associated protein-1 in neuronal cells implicates a role in neuritic growth. Mol Cell Neurosci 16:168-183[Medline]
Li X, Li S, Sharp A, Nucifora F, Schilling G, Lanahan A, Worley P, Snyder S et al. (1995) A huntingtin-associated protein enriched in brain with implications for pathology. Nature 378:398-402[Medline]
Li Y, Chin LS, Levey AI, Li L (2002) Huntingtin-associated protein 1 interacts with hepatocyte growth factor-regulated tyrosine kinase substrate and functions in endosomal trafficking. J Biol Chem 277:28212-28221
Lin BZ, Pilch PF, Kandror KV (1997) Sortilin is a major protein component of Glut4-containing vesicles. J Biol Chem 272:24145-24147
Martin E, Kim M, Velier J, Sapp E, Lee H, Laforet G, Won L et al. (1999) Analysis of huntingtin-associated protein 1 in mouse brain and immortalized striatal neurons. J Comp Neurol 403:421-430[Medline]
Mazella J, Zsurger N, Navarro V, Chabry J, Kaghad M, Caput D, Ferrara P et al. (1998) The 100-kDa neurotensin receptor is gp95/sortilin, a non-G-protein-coupled receptor. J Biol Chem 273:26273-26276
Motoi Y, Aizama T, Haga S, Nakumura S, Namba Y, Ikeda K (1999) Neuronal localization of a novel mosaic apolipoprotein E receptor, LR11, in rat and human brain. Brain Res 833:209-215[Medline]
Navarro A, Rey C, Gd Tolivia J, AlvarezUria M (1996) Ultrastructural and quantitative study of atypical age-related bodies in the hamster brain. Mech Ageing Dev 90:75-90[Medline]
Nielsen MS, Madsen P, Christensen EI, Nykjaer A, Gliemann J, Kasper D, Pohlmann R et al. (2001) The sortilin cytoplasmic tail conveys Golgi-endosome transport and binds the VHS domain of the GGA2 sorting protein. EMBO J 20:2180-2190
Paxinos G, Watson C (1986) The Rat Brain in Stereotaxic Coordinates. 2nd ed San Diego, Academic Press, Harcourt Brace Jovanovich
Pelaez AM, AlvarezUria M (1987) Nucleolus-like bodies in the posterior subnucleus of the rat paraventricular nucleus during postnatal development. An ultrastructural, cytochemical and morphometric study. J Submicrosc Cytol 19:101-105[Medline]
Petersen CM, Nielsen MS, Nykjaer A, Jacobsen L, Tommerup N, Rasmussen HH, Roigaard H et al. (1997) Molecular identification of a novel candidate sorting receptor purified from human brain by receptor-associated protein affinity chromatography. J Biol Chem 272:3599-3605
Qu D, Teckman JH, Omura S, Perlmutter DH (1996) Degradation of a mutant secretory protein, alpha1-antitrypsin Z, in the endoplasmic reticulum requires proteasome activity. J Biol Chem 271:22791-22795
RosenbergHasson Y, Bercovich Z, Ciechanover A, Kahana C (1989) Degradation of ornithine decarboxylase in mammalian cells is ATP dependent but ubiquitin independent. Eur J Biochem 185:469-474[Abstract]
Santolaya R (1973) Nucleolus-like bodies in the neuronal cytoplasm of the mouse arcuate nucleus. Z Zellforsch 146:319-328[Medline]
Serrando M, Casanovas A, Esquerda JE (2002) Occurrence of glutamate receptor subunit 1-containing aggresome-like structures during normal development of rat spinal cord interneurons. J Comp Neurol 442:23-34[Medline]
Shimizu N, Ishii S (1965) Electron microscopic observations on nucleolar extrusion in nerve cells of the rat hypothalamus. Z Zellforsch 67:367-372[Medline]
Shinoda K, Nagano M, Osawa Y (1993) An aromatase-associated cytoplasmic inclusion, the "stigmoid body," in the rat brain: II. Ultrastructure (with a review of its history and nomenclature). J Comp Neurol 329:1-19[Medline]
Taira K, Bujo H, Hirayama S, Yamazaki H, Kanaki T, Takahashi K, Ishii I et al. (2001) LR11, a mosaic LDL receptor family member, mediates the uptake of ApoE-rich lipoproteins in vitro. Arterioscler Thromb Vasc Biol 21:1501-1506
Tasso F, Rua S (1978) Ultrastructural observations on the hypothalamo-posthypophysial complex of the Brattleboro rat. Cell Tissue Res 191:267-286[Medline]
Ward CL, Omura S, Kopito RR (1995) Degradation of CFTR by the ubiquitin-proteasome pathway. Cell 83:121-127[Medline]
Weakley B (1969) Granular cytoplasmic bodies in oocytes of the golden hamster during the postnatal period. Z Zellforsch 101:394-400[Medline]
Wiertz EJ, Jones TR, Sun L, Bogyo M, Geuze HJ, Ploegh HL (1996a) The human cytomegalovirus US11 gene product dislocates MHC class I heavy chains from the endoplasmic reticulum to the cytosol. Cell 84:769-779[Medline]
Wiertz EJ, Tortorella D, Bogyo M, Yu J, Mothes W, Jones TR, Rapoport TA et al. (1996b) Sec61-mediated transfer of a membrane protein from the endoplasmic reticulum to the proteasome for destruction. Nature 384:432-438[Medline]
Yamazaki H, Bujo H, Kusunoki J, Seimiya K, Kanaki T, Morisaki N, Schneider WJ et al. (1996) Elements of neural adhesion molecules and a yeast vacuolar protein sorting receptor are present in a novel mammalian low density lipoprotein receptor family member. J Biol Chem 271:24761-24768
Yamazaki H, Bujo H, Saito Y (1997) A novel member of the LDL receptor gene family with eleven binding repeats is structurally related to neural adhesion molecules and a yeast vacuolar protein sorting receptor. J Atheroscler Thromb 4:20-26[Medline]
Zhu Y, Bujo H, Yamazaki H, Hirayama S, Kanaki T, Takahashi K, Shibasaki M et al. (2002) Enhanced expression of the LDL receptor family member LR11 increases migration of smooth muscle cells in vitro. Circulation 105:1830-1836