Glycolytic enzymes and assembly of microtubule networks

Völker, Katharina; Reinitz, Catharine A.; Knull, Harvey R.

doi:10.1016/0305-0491(95)00096-8

Cited by 63 publications

(36 citation statements)

References 35 publications

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“…Numerous studies have demonstrated interactions between the cytoskeleton and glycolytic enzymes, especially GAPDH, aldolase and PK. [25][26][27][28][29][30][31][32][33][34][35][36][37][38] In this study, quantities of these glycolytic enzymes, as well as actin, tubulin and tropomyosin, were sufficient in U87 pseudopodia to be detected with Figure 4 DeCyder ratios Coomassie blue staining, thus establishing the potential for the energy-generating portion of the glycolytic pathway to energize cytoskeletal proteins for migration within U87 pseudopodia. Polymerization of actin and tubulin shares a similar use of energy from nucleotide triphosphate hydrolysis.…”

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confidence: 65%

Proteomic characterization of harvested pseudopodia with differential gel electrophoresis and specific antibodies

Beckner

Chen

et al. 2005

Laboratory Investigation

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Malignant gliomas (astrocytomas) are lethal tumors that invade the brain. Invasive cell migration is initiated by extension of pseudopodia into interstitial spaces. In this study, U87 glioma cells formed pseudopodia in vitro as cells pushed through 3 lm pores of polycarbonate membranes. Harvesting pseudopodia in a novel two-step method provided material for proteomic analysis. Differences in the protein profiles of pseudopodia and whole cells were found using differential gel electrophoresis (DIGE) and immunoblotting. Proteins from twodimensional (2D) gels with M R 's of 20-100 kDa and pI's of 3.0-10.0 were identified by peptide mass fingerprinting analysis using mass spectrometry. For DIGE, lysates of pseudopodia and whole cells were each labeled with electrophilic forms of fluorescent dyes, Cy3 or Cy5, and analyzed as mixtures. Analysis was repeated with reciprocal labeling. Differences in protein distributions were detected by manual inspection and computer analysis. Topographical digital maps of the scanned gels were used for algorithmic spot matching, normalization of background, quantifying spot differences, and elimination of artifacts. Pseudopodial proteins in Coomassie-stained 2D gels included isoforms of glycolytic enzymes as the largest group, seven of 24 proteins. Peptide mass fingerprint analysis of DIGE gels demonstrated increased isoforms of annexin (Anx) I, AnxII, enolase, pyruvate kinase, and aldolase, and decreased mitochondrial manganese superoxide dismutase and transketolase in pseudopodia. Specific antibodies showed restricted immunoreactivity of the hepatocyte growth factor (HGF) a chain to pseudopodia, indicating localization of its active form. Met (the HGF receptor), actin, and total AnxI were increased in pseudopodial lysates on immunoblots. Increased constituents of the pseudopodial proteome in glioma cells, identified in this study as actin, HGF, Met, and isoforms of AnxI, AnxII, and several glycolytic enzymes, represent therapeutic targets to consider for suppression of tumor invasion.

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confidence: 65%

Proteomic characterization of harvested pseudopodia with differential gel electrophoresis and specific antibodies

Beckner

Chen

et al. 2005

Laboratory Investigation

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“…Most glycolytic enzymes, including PGAM, exist in complexes bound to the microtubule network (26,27). The GTPases Rac and Cdc42 regulate not only F-actin reorganization but also microtubule dynamics through Pak, which phosphorylates and thereby inhibits the microtubule-destabilizing protein stathmin, leading to microtubule stabilization (28).…”

Section: Discussionmentioning

confidence: 99%

A p21-Activated Kinase-controlled Metabolic Switch Up-regulates Phagocyte NADPH Oxidase

Shalom‐Barak

Knaus

2002

Journal of Biological Chemistry

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Chemoattractant-stimulated phagocytes increase their glucose uptake and divert energy production from glycolysis to the pentose phosphate pathway to generate NADPH. NADPH is a required cofactor for the NADPH oxidase to produce reactive oxygen metabolites, an important microbicidal tool in host defense. p21-Activated kinases (Paks) are regulated by the GTPases Rac and Cdc42 and control actin dynamics and phosphorylation of the oxidase component p47phox . Here we report the interaction of Pak with phosphoglycerate mutase (PGAM)-B, an enzyme of the glycolytic pathway. Activated Pak1 inhibits glycolysis by association of its catalytic domain with PGAM-B and subsequent phosphorylation of the enzyme on serine residues 23 and 118, thereby abolishing PGAM activity. Leukocyte activation through chemoattractant receptors leads to Pak activation and transient inhibition of endogenous PGAM-B activity. Consistent with these observations, treatment of neutrophils with phosphoglycolic acid, a competitive PGAM-B inhibitor, increases upstream intermediates, thereby amplifying the respiratory burst. These results demonstrate that Rho GTPases regulate the glycolytic pathway through Pak and suggest a link between chemoattractant signaling and metabolic responses to enhance host defense.The invasion of pathogens presents an acute challenge to the host organism, which requires an immediate defensive response. Innate immune cells, mainly macrophages, neutrophils, Kupffer cells, and microglia, are specialized in the physical destruction of the invading pathogen via phagocytosis and oxidative burst. These activities are elicited by the binding of chemoattractants including bacterial-derived formylated peptides (formyl-Met-Leu-Phe) to G protein-coupled receptors on the cell surface. The chemotactic stimulus leads to activation of the low molecular weight GTP-binding proteins Rac and Cdc42 and their downstream mediators p21-activated kinases (Paks) 1 (1, 2). Rac and Pak signaling is essential in cytoskeletal reorganization including membrane ruffling and formation of filopodia and lamellipodia, thus enabling cell motility during the chemotactic response (3-5), as well as in pathogen-induced respiratory burst activity mediated by a Rac2-regulated multimeric NADPH oxidase enzyme (2, 6). The NADPH oxidase converts molecular oxygen to superoxide anion by electron transport from NADPH, a process that involves translocation and association of active Rac, p67phox , and phosphorylated p47 phox with membrane-bound cytochrome b 558 (7). Pak participates in this pathway by phosphorylation of several serine residues on p47 phox , which are required for oxidase activity and possibly for assembly of the active oxidase (8). Neutrophils contain at least three Pak isoforms whose fMLF-induced activation kinetics coincide with Rac2 activation, p47 phox phosphorylation, and oxidant production (8, 9).Uptake and destruction of pathogens during phagocytosis are accompanied by simultaneous elevation of glucose transporter 1 translocation, glucose uptake, and oxy...

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“…It is possible that this interaction depends on appropriate modifications or/and physiological conditions such as phosphorylation or/ and Ca 2+ concentration (19)(20)(21). It has been proposed that in the cytoplasm of tumor cells, ·-enolase and other glycolytic enzymes interact with the cytoskeleton system and that such association may provide ATP to promote the migration of tumor cells (22)(23)(24). Furthermore, ·-enolase can also be expressed on the cell surface depending on the activation status or pathophysiological conditions (25).…”

Section: Discussionmentioning

confidence: 99%

Expression of the sperm fibrous sheath protein CABYR in human cancers and identification of α-enolase as an interacting partner of CABYR-a

Yang

2011

Oncol Rep

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Glycolytic enzymes and assembly of microtubule networks

Cited by 63 publications

References 35 publications

Proteomic characterization of harvested pseudopodia with differential gel electrophoresis and specific antibodies

Proteomic characterization of harvested pseudopodia with differential gel electrophoresis and specific antibodies

A p21-Activated Kinase-controlled Metabolic Switch Up-regulates Phagocyte NADPH Oxidase

Expression of the sperm fibrous sheath protein CABYR in human cancers and identification of α-enolase as an interacting partner of CABYR-a

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