Biomedical subjects

C M Petersen

Publications and source records attributed to C M Petersen.

At least 19 recordsLinked to original sources

PubMed2005

SorCS3 does not require propeptide cleavage to bind nerve growth factor.

The functional properties of the Vps10p-domain receptor SorCS3 are undescribed. Here, we examine its processing and sorting in cellular transfectants, and analyze the binding of potential ligands to the purified receptor. We show that SorCS3 is synthesized as a proprotein and converted to its mature form by N-terminal propeptide cleavage in distal Golgi compartments. The propeptide is not a requirement for normal processing of the receptor and does not prevent ligands from binding to the SorCS3 precursor form. Expression of wt and chimeric receptors further suggests that SorCS3 predominates on the plasma membrane, exhibits slow internalization and does not engage in intracellular trafficking. SorCS3 emerges as a new neurotrophin binding Vps10p-domain receptor functionally distinct from its relatives Sortilin and SorLA.

Animals↗

PubMed2001

The sortilin cytoplasmic tail conveys Golgi-endosome transport and binds the VHS domain of the GGA2 sorting protein.

Sortilin belongs to a growing family of multiligand type-1 receptors with homology to the yeast receptor Vps10p. Based on structural features and sortilin's intracellular predominance, we have proposed it to be a sorting receptor for ligands in the synthetic pathway as well as on the cell membrane. To test this hypothesis we examine here the cellular trafficking of chimeric receptors containing constructs of the sortilin tail. We report that sorting signals conforming to YXX and dileucine motifs mediate rapid endocytosis of sortilin chimeras, which subsequently travel to the trans-Golgi network, showing little or no recycling. Furthermore, we found that cation-independent mannose 6-phosphate receptor (MPR300)-sortilin chimeras, expressed in mannose 6-phosphate receptor knockout cells, were almost as efficient as MPR300 itself for transport of newly synthesized beta-hexosaminidase and beta-glucuronidase to lysosomes, and established that the sortilin tail contains potent signals for Golgi-endosome sorting. Finally, we provide evidence suggesting that sortilin is the first example of a mammalian receptor targeted by the recently described GGA family of cytosolic sorting proteins, which condition the Vps10p-mediated sorting of yeast carboxypeptidase Y.

Adaptor Proteins, Vesicular Transport↗

PubMed2001

Pharmacological properties of the mouse neurotensin receptor 3. Maintenance of cell surface receptor during internalization of neurotensin.

We recently reported the molecular identification of a new type of receptor for the neuropeptide neurotensin (NT), the neurotensin receptor 3 (NTR3), identical to sortilin, which binds receptor-associated protein. Here, we demonstrate that the cloned mouse NTR3 is expressed on the plasma membrane of transfected COS-7 cells. The mouse NTR3 is detectable by photoaffinity labeling and immunoblotting at the cell surface as a 100 kDa N-glycosylated protein. Biochemical analysis and confocal microscopic imaging clearly indicate that NT is efficiently internalized after binding to NTR3, and that despite this internalization, the amount of receptor present on the cell surface is maintained.

Adaptor Proteins, Vesicular Transport↗

PubMed2001

Activation and functional characterization of the mosaic receptor SorLA/LR11.

We previously isolated and sequenced the approximately 250-kDa type 1 receptor sorLA/LR11, a mosaic protein with elements characterizing the Vps10p domain receptor family as well as the low density lipoprotein receptor family. The N terminus of the Vps10p domain comprises a consensus sequence for cleavage by furin ((50)RRKR(53)) that precedes a truncation found in sorLA isolated from human brain. Here we show that sorLA, like sortilin-1/neurotensin receptor-3, whose lumenal domain consists of a Vps10p domain only, is synthesized as a proreceptor that is cleaved by furin in late Golgi compartments. We show that the truncation conditions the Vps10p domain for propeptide inhibitable binding of neuropeptides and the receptor-associated protein. We further demonstrate that avid binding of the receptor-associated protein, apolipoprotein E, and lipoprotein lipase not inhibited by propeptide occurs to sites located in other lumenal domains. In transfected cells, about 10% of full-length sorLA were expressed on the cell surface capable of mediating endocytosis. However, the major pool of receptors was found in late Golgi compartments, suggesting possible interaction with newly synthesized ligands. The results show that sorLA, following activation by truncation, binds multiple ligands and may mediate both endocytosis and sorting.

Adaptor Proteins, Vesicular Transport↗

PubMed2001

Reliability of assessing end-feel and pain and resistance sequence in subjects with painful shoulders and knees.

STUDY DESIGN: Descriptive. OBJECTIVES: Examine the intrarater and interrater reliability of end-feel and pain/resistance sequence for patients with painful shoulders and knees. BACKGROUND: Clinicians make diagnostic and intervention decisions based on end-feel and pain/resistance sequence, but few studies have examined agreement within and between physical therapists when assessing subjects with pathology. METHODS AND MEASURES: Subjects with unilateral knee pain (18 men and 22 women with a mean age of 31.8 +/- 9.5 years) or shoulder pain (21 men and 25 women with a mean age of 34.3 +/- 12.9 years) were examined twice. Two physical therapists used standardized positions to evaluate 2 knee motions and 5 shoulder motions. Evaluators did not interview subjects and were blinded to previous test results. Evaluators applied overpressure and noted the end-feel while subjects identified the moment their pain was reproduced. Following testing, subjects rated their pain intensity. Analyses included: percentage of agreement; kappa, weighted kappa, and maximum kappa coefficients; and confidence intervals. Analyses were repeated for subjects whose pain intensity during testing did not change between examinations. RESULTS: Intrarater kappa coefficients varied from 0.65 to 1.00 for end-feel, and intrarater weighted kappa coefficients varied from 0.59 to 0.87 for pain/resistance sequence. Most coefficients remained stable or improved for the unchanged subjects. Interrater kappa coefficients for end-feel and weighted kappa coefficients for pain/resistance sequence varied from -0.01 to 0.70. End-feel kappa coefficients remained low for the unchanged subjects, but pain/resistance sequence weighted kappa coefficients improved. Unbalanced distribution affected many coefficients, producing low coefficients even when the percentage of agreement was high. CONCLUSIONS: The appropriate use of end-feel and pain/resistance sequence data requires reliable data gathering, especially when patients are managed by more than one physical therapist. Intrarater reliability of end-feel and pain/resistance judgments at the knee and shoulder were generally good, especially after accounting for subject change and unbalanced distributions. Interrater reliability, however, was generally not acceptable, even after accounting for these factors.

Adult↗

PubMed2000

Reliability of cervical range of motion using the OSI CA 6000 spine motion analyser on asymptomatic and symptomatic subjects.

Cervical range of motion (ROM) is evaluated in both clinical and research settings. This study's purpose was to determine if ROM data obtained with the OSI CA 6000 Spine Motion Analyser (SMA) from asymptomatic and symptomatic cervical subjects were reliable within and between testers. Cervical ROM was measured in all three planes in 30 adult asymptomatic and 20 adult symptomatic subjects. A standardized protocol was used to fit each subject with the OSI SMA cervical hardware. Subjects were tested in a seated position with the trunk stabilized. Subjects performed four trials of each pain-free cervical motion during testing. The hardware was completely removed and replaced by the same tester and ROM trials in all three planes were repeated for intratester asymptomatic and symptomatic reliability. The same procedure was completed by a second tester for asymptomatic intratester and intertester reliability. Repeated measures analysis of variance and intraclass correlation coefficients (ICC [2,1 and 2 k]) were used to analyse intra- and intertester reliability data. Intratester ICCs were 0.85 or higher (except for flexion 0.76) for asymptomatic subjects and 0. 87 or higher (except for flexion 0.68) for symptomatic subjects for all motions. Intertester ICCs were 0.88 or higher for all motions. Standard error of measurements were less than 3.92 degrees for all motions. Measures of cervical spinal ROM obtained with the OSI SMA showed good intertester reliablity for all motions, and good intratester reliability for all motions with the exception of the motion of flexion for one of the examiners, which showed moderate reliability.

Adult↗

PubMed2000

Construct validity of Cyriax's selective tension examination: association of end-feels with pain at the knee and shoulder.

STUDY DESIGN: Descriptive. OBJECTIVES: To examine the relationship between pain and normal and abnormal-pathologic end-feels during passive physiologic motion assessment at the knee and shoulder. We theorized that abnormal-pathologic end-feels would be more painful than normal end-feels. BACKGROUND: End-feel testing and pain intensity information are part of physical therapy musculoskeletal patient examinations. End-feels are categorized as normal or abnormal-pathologic. No previous studies have examined the relationship between pain during end-feel testing and the type of end-feel. METHODS AND MEASURES: Two physical therapists examined subjects with unilateral knee or shoulder pain. Each subject was examined twice. Passive physiologic motions, 2 at the knee and 5 at the shoulder, were tested by applying an overpressure at the end of range of motion using standardized positions. Subjects reported the amount of pain (0-10) immediately after the evaluator recorded the end-feel. Analyses included one-way ANOVAs and post-hoc Tukey's Honestly Significant Difference tests. RESULTS: Some abnormal-pathologic end-feels were significantly more painful than the normal end-feels at both the knee and the shoulder for all physiologic motions. Among the abnormal-pathologic end-feel categories there were no statistical differences in pain intensity, although small samples in some categories may be responsible for this finding. CONCLUSION: Abnormal-pathologic end-feels are associated with more pain than normal end-feels during passive physiologic motion testing at the knee or shoulder. Dysfunction should be suspected when abnormal-pathologic end-feels are present.

Adult↗

PubMed1999

Sortilin/neurotensin receptor-3 binds and mediates degradation of lipoprotein lipase.

Lipoprotein lipase and the receptor-associated protein (RAP) bind to overlapping sites on the low density lipoprotein receptor-related protein/alpha2-macroglobulin receptor (LRP). We have investigated if lipoprotein lipase interacts with the RAP binding but structurally distinct receptor sortilin/neurotensin receptor-3. We show, by chemical cross-linking and surface plasmon resonance analysis, that soluble sortilin binds lipoprotein lipase with an affinity similar to that of LRP. The binding was inhibited by heparin and RAP and by the newly discovered sortilin ligand neurotensin. In 35S-labeled 3T3-L1 adipocytes treated with the cross-linker dithiobis(succinimidyl propionate), lipoprotein lipase-containing complexes were isolated by anti-sortilin antibodies. To elucidate function in cells, sortilin-negative Chinese hamster ovary cells were transfected with full-length sortilin and shown to express about 8% of the receptors on the cell surface. These cells degraded 125I-labeled lipoprotein lipase much faster than the wild-type cells. The degradation was inhibited by unlabeled lipoprotein lipase, indicating a saturable pathway, and by RAP and heparin. Moreover, inhibition by the weak base chloroquine suggested that degradation occurs in an acidic vesicle compartment. The results demonstrate that sortilin is a multifunctional receptor that binds lipoprotein lipase and, when expressed on the cell surface, mediates its endocytosis and degradation.

Adaptor Proteins, Vesicular Transport↗

PubMed1999

A head-activator binding protein is present in hydra in a soluble and a membrane-anchored form.

The neuropeptide head activator plays an important role for proliferation and determination of stem cells in hydra. By affinity chromatography a 200 kDa head-activator binding protein, HAB, was isolated from the multiheaded mutant of Chlorohydra viridissima. Partial amino acid sequences were used to clone the HAB cDNA which coded for a receptor with a unique alignment of extracellular modules, a transmembrane domain, and a short carboxy-terminal cytoplasmic tail. A mammalian HAB homologue with identical alignment of these modules is expressed early in brain development. Specific antibodies revealed the presence of HAB in hydra as a transmembrane receptor, but also as secreted protein, both capable of binding head activator. Secretion of HAB during regeneration and expression in regions of high determination potential hint at a role for HAB in regulating the concentration and range of action of head activator.

Amino Acid Sequence↗

PubMed1998

The carboxy-terminal domain of the receptor-associated protein binds to the Vps10p domain of sortilin.

Binding of the receptor-associated protein (RAP) to the newly identified putative sorting receptor, sortilin, was analyzed by surface plasmon resonance analysis of recombinant RAP and sortilin domains and compared with binding to megalin and low density lipoprotein receptor-related protein (LRP). The data show that the RAP-binding site in sortilin is localized in the cysteine-rich lumenal part homologous to yeast vacuolar protein-sorting 10 protein (Vps10p), and the sortilin-binding site in RAP is localized in the carboxy-terminal domain III of the three homologous domains in RAP. Whereas sortilin bound only RAP domain III, megalin and LRP bound all RAP domains with the functional affinity order: domain III >domain I > domain II.

Adaptor Proteins, Vesicular Transport↗

PubMed1998

Mannose 6-phosphate/insulin-like growth factor-II receptor targets the urokinase receptor to lysosomes via a novel binding interaction.

The urokinase-type plasminogen activator receptor (uPAR) plays an important role on the cell surface in mediating extracellular degradative processes and formation of active TGF-beta, and in nonproteolytic events such as cell adhesion, migration, and transmembrane signaling. We have searched for mechanisms that determine the cellular location of uPAR and may participate in its disposal. When using purified receptor preparations, we find that uPAR binds to the cation-independent, mannose 6-phosphate/insulin-like growth factor-II (IGF-II) receptor (CIMPR) with an affinity in the low micromolar range, but not to the 46-kD, cation-dependent, mannose 6-phosphate receptor (CDMPR). The binding is not perturbed by uPA and appears to involve domains DII + DIII of the uPAR protein moiety, but not the glycosylphosphatidylinositol anchor. The binding occurs at site(s) on the CIMPR different from those engaged in binding of mannose 6-phosphate epitopes or IGF-II. To evaluate the significance of the binding, immunofluorescence and immunoelectron microscopy studies were performed in transfected cells, and the results show that wild-type CIMPR, but not CIMPR lacking an intact sorting signal, modulates the subcellular distribution of uPAR and is capable of directing it to lysosomes. We conclude that a site within CIMPR, distinct from its previously known ligand binding sites, binds uPAR and modulates its subcellular distribution.

Amino Acid Sequence↗

PubMed1998

Sortilin is the major 110-kDa protein in GLUT4 vesicles from adipocytes.

Vesicles containing the glucose transporter GLUT4 from rat adipocytes contain a major protein of 110 kDa. We have isolated this protein, obtained the sequences of peptides, and cloned a large portion of its cDNA. This revealed that the protein is sortilin, a novel membrane protein that was cloned in another context from a human source while this work was in progress. Subcellular fractionation of rat and 3T3-L1 adipocytes, together with GLUT4 vesicle isolation, showed that sortilin was primarily located in the low density microsomes in vesicles containing GLUT4. Insulin caused a 1.7-fold increase in the amount of sortilin at the plasma membranes of 3T3-L1 adipocytes, as assessed by cell surface biotinylation. The expression of sortilin in 3T3-L1 cells occurred only upon differentiation. Previous characterization of sortilin has led to the suggestion that it functions to sort lumenal proteins from the trans Golgi. The significance of its insulin-stimulated increase at the cell surface and of its expression upon differentiation will require definitive delineation of its function.

3T3 Cells↗

PubMed1997

Molecular identification of a novel candidate sorting receptor purified from human brain by receptor-associated protein affinity chromatography.

Receptor-associated protein (RAP) is an endoplasmic reticulum/Golgi protein involved in the processing of receptors of the low density lipoprotein receptor family. A approximately 95-kDa membrane glycoprotein, designated gp95/sortilin, was purified from human brain extracts by RAP affinity chromatography and cloned in a human cDNA library. The gene maps to chromosome 1p and encodes an 833-amino acid type I receptor containing an N-terminal furin cleavage site immediately preceding the N terminus determined in the purified protein. Gp95/sortilin is expressed in several tissues including brain, spinal cord, and testis. Gp95/sortilin is not related to the low density lipoprotein receptor family but shows intriguing homologies to established sorting receptors: a 140-amino acid lumenal segment of sortilin representing a hitherto unrecognized type of extracellular module shows extensive homology to corresponding segments in each of the two lumenal domains of yeast Vps10p, and the extreme C terminus of the cytoplasmic tail of sortilin contains the casein kinase phosphorylation consensus site and an adjacent dileucine sorting motif that mediate assembly protein-1 binding and lysosomal sorting of the mannose-6-phosphate receptors. Expression of a chimeric receptor containing the cytoplasmic tail of gp95/sortilin demonstrates evidence that the tail conveys colocalization with the cation-independent mannose6-phosphate receptor in endosomes and the Golgi compartment.

Adaptor Proteins, Vesicular Transport↗

PubMed1996

Molecular characterization of a novel human hybrid-type receptor that binds the alpha2-macroglobulin receptor-associated protein.

The 39-40-kDa receptor-associated protein (RAP) binds to the members of the low density lipoprotein receptor gene family and functions as a specialized endoplasmic reticulum/Golgi chaperone. Using RAP affinity chromatography, we have purified a novel approximately 250-kDa brain protein and isolated the corresponding cDNA. The gene, designated SORL1, maps to chromosome 11q 23/24 and encodes a 2214-residue type 1 receptor containing a furin cleavage site immediately preceding the N terminus determined in the purified protein. The receptor, designated sorLA-1, has a short cytoplasmic tail containing a tyrosine-based internalization signal and a large external part containing (from the N-terminal): 1) a segment homologous to domains in the yeast vacuolar protein sorting 10 protein, Vps10p, that binds carboxypeptidase Y, 2) five tandemly arranged YWTD repeats and a cluster of 11 class A repeats characteristic of the low density lipoprotein receptor gene family receptors, and 3) six tandemly arranged fibronectin type III repeats also found in certain neural adhesion proteins. sorLA-1 may therefore be classified as a hybrid receptor. Northern blotting revealed specific mRNA transcripts in brain, spinal cord, and testis but not in several major organs. Both RAP and an antibody against a synthetic peptide derived from a sequence determined in the mature protein detected sorLA-1 in crude human brain extracts. The domain structure suggests that sorLA-1 is an endocytic receptor possibly implicated in the uptake of lipoproteins and of proteases.

Amino Acid Sequence↗

PubMed1996

The receptor-associated protein (RAP) binds calmodulin and is phosphorylated by calmodulin-dependent kinase II.

The receptor-associated protein, RAP, is an intracellular protein that may function as a chaperone for the LDL-receptor family receptors. Here we report calmodulin as the first identified RAP binding protein outside of the LDL-receptor family members. We demonstrate that RAP binds calmodulin in a Ca2+- and pH-dependent manner characteristic of calmodulin-dependent enzymes, and present evidence that RAP is a substrate for calmodulin-dependent enzymes. Thus, CaM-kinase II and calcineurin readily phosphorylate and dephosphorylate, respectively, serine residues in RAP, and in the individual RAP domains D2 (amino acids 113-218) and D3 (amino acids 219-323) which both contain sites for CaM-kinase II-mediated phosphorylation and for calmodulin binding. In addition, we provide evidence that RAP is phosphorylated by other kinases such as casein kinase II. Studies of 32[ortho]P-labelled cell cultures demonstrate that RAP is phosphorylated in vivo. Our results suggest that RAP may have hitherto unknown functions implicating phosphorylation and calmodulin-mediated modulation.

Amino Acid Sequence↗

PubMed1996

Megalin-mediated endocytosis of transcobalamin-vitamin-B12 complexes suggests a role of the receptor in vitamin-B12 homeostasis.

Kidney cortex is a main target for circulating vitamin B12 (cobalamin) in complex with transcobalamin (TC). Ligand blotting of rabbit kidney cortex with rabbit 125I-TC-B12 and human TC-57Co-B12 revealed an exclusive binding to megalin, a 600-kDa endocytic receptor present in renal proximal tubule epithelium and other absorptive epithelia. The binding was Ca2+ dependent and inhibited by receptor-associated protein (RAP). Surface plasmon resonance analysis demonstrated a high-affinity interaction between purified rabbit megalin and rabbit TC-B12 but no measurable affinity of the vitamin complex for the homologous alpha 2-macroglobulin receptor (alpha 2MR)/low density lipoprotein receptor related protein (LRP). 125I-TC-B12 was efficiently endocytosed in a RAP-inhibitable manner in megalin-expressing rat yolk sac carcinoma cells and in vivo microperfused rat proximal tubules. The radioactivity in the tubules localized to the endocytic compartments and a similar apical distribution in the proximal tubules was demonstrated after intravenous injection of 125I-TC-B12. The TC-B12 binding sites in the proximal tubule epithelium colocalized with megalin as shown by ligand binding to cryosections of rat kidney cortex, and the binding was inhibited by anti-megalin polyclonal antibody, EDTA, and RAP. These data show a novel nutritional dimension of megalin as a receptor involved in the cellular uptake of vitamin B12. The expression of megalin in absorptive epithelia in the kidney and other tissues including yolk sac and placenta suggests a role of the receptor in vitamin B12 homeostasis and fetal vitamin B12 supply.

Animals↗

PubMed1996

Cloning and sequencing of a human cDNA encoding a putative transcription factor containing a bromodomain.

A 2985 bp cDNA was isolated from a Lambda Zap Express library and sequenced. The cDNA appeared to represent a previously unknown gene that encodes and acidic 757 amino acid protein containing a bromodomain, several potential sites for phosphorylation by casein kinase-II and small proline-rich segments. The results suggest that the encoded protein might be a novel transcription factor.

Amino Acid Sequence↗

PubMed1996

Soluble tumor necrosis factor (TNF) receptors conserve TNF bioactivity in meningitis patient spinal fluid.

The content of tumor necrosis factor (TNF)-alpha antigen and the bioactivity of soluble TNF receptor type II (sTNF-RII) in cerebrospinal fluid (CSF) from 29 patients with meningeal symptoms and fever were examined. Immunoreactive TNF was demonstrated in CSF from 4 of 7 patients with bacterial meningitis. In 3 of 8 patients with aseptic meningitis, CSF also contained TNF, but TNF bioactivity was confined to samples from patients with bacterial meningitis. Bioactive TNF was exclusively in high-performance liquid chromatography fractions containing 30- to 60-kDa proteins. Lipopolysaccharide induced down-regulation, possibly after shedding of granulocyte surface membrane TNF-RII. Consistently, there was a statistically significant correlation between sTNF-RII and CSF leukocyte counts. Bioactive TNF was found only in CSF containing >1 ng of sTNF-RII/mL; samples without TNF bioactivity contained less sTNF-RII. Thus, a stabilizing effect of sTNF-RII on the oligomeric cytokine in vivo is plausible.

Cells, Cultured↗