Dissecting mannose 6-phosphate-insulin-like growth factor 2 receptor complexes that control activation and uptake of plasminogen in cells

The plasminogen (Plg) activation cascade on the cell surface plays a central role in cell migration and is involved in a plethora of physiological and pathological processes. Its regulation is coordinated by many receptors, in particular the urokinase-type plasminogen activator receptor (uPAR, CD87), receptors that physically interact and functionally cooperate with uPAR, and Plg binding molecules. Here we studied the impact of one of the Plg binding molecules, the mannose 6-phosphate/insulin-like growth factor 2 receptor (M6P-IGF2R, CD222), on cellular Plg activation. By developing both in vitro and in vivo Plg activation assays on size-fractionated lysates of M6P-IGF2R-silenced cells, we identified Plg-associated complexes with M6P-IGF2R as the regulatory factor. Using lipid raft preserving versus dissolving detergents, we found lipid dependence of the Plg regulatory function of these complexes. Furthermore, M6P-IGF2R-silencing in uPAR-positive human cell lines reduced internalization of Plg, resulting in elevated Plg activation. In contrast, the expression of human M6P-IGF2R in mouse embryonic fibroblasts derived from M6P-IGF2R knock-out mice enhanced Plg internalization. Finally, peptide 18-36 derived from the Plg-binding site within M6P-IGF2R enhanced Plg uptake. Thus, by targeting Plg to endocytic pathways, M6P-IGF2R appears to control Plg activation within cells that might be important to restrict plasmin activity to specific sites and substrates.     

Mannose 6-Phosphate/Insulin-like Growth Factor 2 Receptor Limits Cell Invasion by Controlling αVβ3 Integrin Expression and Proteolytic Processing of Urokinase-type Plasminogen Activator Receptor

The multifunctional mannose 6-phosphate/insulin-like growth factor 2 receptor (M6P/IGF2R) is considered a tumor suppressor. We report here that RNA interference with M6P/IGF2R expression in urokinase-type plasminogen activator (uPA)/urokinase-type plasminogen activator receptor (uPAR) expressing human cancer and endothelial cells resulted in increased pericellular plasminogen activation, cell adhesion, and higher invasive potential through matrigel. M6P/IGF2R silencing led also to the cell surface accumulation of urokinase and plasminogen and enhanced expression of αV integrins. Genetic rescue experiments and inhibitor studies revealed that the enhanced plasminogen activation was due to a direct effect of M6P/IGF2R on uPAR, whereas increased cell adhesion to vitronectin was dependent on αV integrin expression and not uPAR. Increased cell invasion of M6P/IGF2R knockdown cells was rescued by cosilencing both uPAR and αV integrin. Furthermore, we found that M6P/IGF2R expression accelerates the cleavage of uPAR. M6P/IGF2R silencing resulted in an increased ratio of full-length uPAR to the truncated D2D3 fragment, incapable of binding most uPAR ligands. We conclude that M6P/IGF2R controls cell invasion by regulating αV integrin expression and by accelerating uPAR cleavage, leading to the loss of the urokinase/vitronectin/integrin-binding site on uPAR.     

Mineralization of 3-chloro-4-methylaniline via an ortho-cleavage pathway by Pseudomonas cepacia strain CMA1

Pseudomonas cepacia strain CMA1, which was isolated from soil, utilized 3-chloro-4-methylaniline (3C4MA) in concentrations up to 1.4 mm (0.2 g·l^–1) as the sole source of carbon, nitrogen, and energy. In addition, 3-chloroaniline, 4-chloroaniline and phenol, but not aniline or methylanilines, were degraded by strain CMA1. Biodegradation of the anilines was coupled to the liberation of ammonium and chloride. The broad specificities of the aniline- and catechol-oxidizing enzymes were demonstrated in oxygen uptake experiments, which in addition showed higher activities for ring-cleaving than for aniline-oxidizing enzymes. Two ring-cleaving catechol 1,2-dioxygenases, which were induced selectively after growth on 3C4MA (pyrocatechase type II) and phenol (pyrocatechase type I), respectively, were discerned after partial purification by DEAE-cellulose chromatography. Correspondence to: F. Streichsbier     

Molecular characterization and functional analysis of the leukocyte surface protein CD31.

The CD31 Ag is a surface glycoprotein of 130 kDa with a broad cellular distribution. We show that among peripheral human blood cells, it is expressed on monocytes, granulocytes, platelets, and a subpopulation of lymphocytes. Activation of granulocytes leads to down-regulation of CD31 molecule expression. Sequence analysis and quantitative measurements of the relatedness of the CD31 molecule to other known proteins demonstrate that it consists of six Ig constant domains and that each domain bears substantial similarity to Fc gamma R domains. We find, however, that the CD31 molecule does not bind Ig Fc domains. On human monocytes we demonstrate that CD31 mAb recognizing certain epitopes of the CD31 molecule induce the generation of reactive oxygen metabolites. No such effect was seen with human granulocytes. By using two CD31 mAb, termed 1B5 and 7E4, we analyzed the requirements for activation of the monocyte respiratory burst via CD31 Ag in more detail. We show that signal transduction occurs via formation of a CD31 Ag-mAb-Fc gamma R complex involving either Fc gamma RI (CD64) or Fc gamma RII (CDw32) molecules.     

Die Feinstruktur des Glomus caroticum beim Menschen

Zusammenfassung Die Feinstruktur des Glomus caroticum des Menschen wird beschrieben (Operationsmaterial). Die disseminierten Parenchymzellhaufen bestehen aus Glomuszellen und Stützzellen. Die Glomuszellen zeigen alle Kriterien chromaffiner Elemente, die Stützzellen entsprechen strukturell den Schwannschen Zellen. Die fenestrierten Endothelien der Gefäße weisen auf einen regen Stoffaustausch hin. Direkte Kontakte zwischen Glomuszellen und Gefäßen kommen nicht vor. Das nervenreiche Bindegewebe um die Glomuszellhaufen wird als Stratum nervosum bezeichnet. Die Nerven endigen z.T. in dem von Bindegewebszellen gekammerten Stratum nervosum, z.T. ziehen sie in das Organparenchym und bilden dort freie Endigungen, einfache Kontaktflächen und typische synaptische Verbindungen mit den Glomuszellen. Erstmals werden Mitochondriensäcke als rezeptorische Endigungen der Nerven und intraaxonale Hohlräume mit geordneten Innenstrukturen beschrieben, deren Funktion unklar ist.

---

The fine structure of the human carotid body

Summary The fine structure of the human carotid body is described. The diffuse conglomerations of cells consist of glomus cells and sustentacular cells. The glomus cells show all criteria of chromaffine elements. The sustentacular cells correspond structurally to the cells of Schwann. The fenestrated endothelial cells of the blood vessels point to an active exchange of metabolic substances. No direct contact between glomus cells and the blood vessels is observed. The connective tissue which is rich in nervous material close to the glomus cells is defined as the stratum nervosum. The nerves partially end in the stratum nervosum, which is segmented by the cells of the connective tissue and on the other hand reach into the cellparenchyma of the carotid body forming free terminal endings, simple areas of contact and typical synapses with the glomus cells. Mitochondrial sacs are described for the first time as nerve receptors. Intraaxonal spaces with an organized structure are described, whose function is not understood.

     

Ein Beitrag zum Problem der Zellform und deren Umwandlung

Zusammenfassung An Hand von Modellversuchen und Experimenten an lebenden Zellen werden jene Faktoren diskutiert, die die Form der Zellen in Gewebekulturen beeinflussen können, wobei besonders auf die Wirkung von Oberflächenkräften hingewiesen wird. Im Rahmen dieser Untersuchungen wurde unter anderem festgestellt, daß spezifische, gegen Gewebsbrei von Hühnerembryonen gerichtete Antisera von Meerschweinchen an Hühnerfibroblasten von Gewebekulturen irreversible Zustandsänderungen auslösen können, die bei höheren Konzentrationen des Serums bis zu einer Zerstörung der Zellmembran und zu einem Ausfließen des Plasmas führen. Auch das Zytoplasma scheint dann irreversibel verändert zu sein. Bei Verwendung geringerer Konzentrationen des Antiserums kommt es primär zu einer kurzen Bewegungsphase der Zelloberfläche und zu einer Abrundung der Zellen. Ähnliche Erscheinungen können auch durch wenig konzentrierte Lösungen von Tannin erzielt werden. Die Versuchsergebnisse ermöglichen verschiedene Schlußfolgerungen hinsichtlich des Zustandekommens von Formveränderungen der Zellen und der Wirkung spezifischer Antikörper.     

The reelin receptor ApoER2 recruits JNK-interacting proteins-1 and -2

Correct positioning of neurons during embryonic development of the brain depends, among other processes, on the proper transmission of the reelin signal into the migrating cells via the interplay of its receptors with cytoplasmic signal transducers. Cellular components of this signaling pathway characterized to date are cell surface receptors for reelin like apolipoprotein E receptor 2 (ApoER2), very low density lipoprotein receptor (VLDLR), and cadherin-related neuronal receptors, and intracellular components like Disabled-1 and the nonreceptor tyrosine kinase Fyn, which bind to the intracellular domains of the ApoER2 and VLDL receptor or of cadherin-related neuronal receptors, respectively. Here we show that ApoER2, but not VLDLR, also binds the family of JNK-interacting proteins (JIPs), which act as molecular scaffolds for the JNK-signaling pathway. The ApoER2 binding domain on JIP-2 does not overlap with the binding sites for MLK3, MKK7, and JNK. These results suggest that ApoER2 is able to assemble a multiprotein complex containing Disabled-1 and JIPs, together with their binding partners, to the cell surface of neurons. This complex might participate in ApoER2-specific reelin signaling and thus would explain the different phenotype of mice lacking the ApoER2 from that of VLDLR-deficient mice.