Co‐localization of glucokinase with actin filaments

Murata, Takeshi; Katagiri, Hideki; Ishihara, Hideharu; Shibasaki, Yoshikazu; Asano, Tomohiko; Toyoda, Yusuke; Pekiner, Bilgehan; Pekiner, Can; Miwa, Ichitomo; Oka, Yoshitomo

doi:10.1016/s0014-5793(97)00253-6

Cited by 28 publications

(21 citation statements)

References 19 publications

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“…Assuming symmetric transport, the ratio of the maximum glucose transport rate to the glucose consumption rate in COS-7 cells was determined as ϳ10 using non-linear regression (27). The lack of response to 0.1 mM glucose is thus consistent with the detection limit of FLIPglu-600 ( Table I) and indicates that in the absence of external glucose supply, cytosolic levels drop to very low concentrations, potentially in the K m range of 50 M for hexokinases present in COS-7 cells (32,33). Additional affinity mutants will be required to resolve cytosolic concentrations below 65 M. Taken together, the results show that in COS-7 cells, cytosolic glucose concentrations can vary over at least two orders of magnitude.…”

Section: Fig 3 In Vivo Characterization Of Flipglu-600supporting

confidence: 56%

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Imaging of the Dynamics of Glucose Uptake in the Cytosol of COS-7 Cells by Fluorescent Nanosensors

Fehr¹,

Lalonde

Lager

et al. 2003

Journal of Biological Chemistry

283

257

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Glucose homeostasis is a function of glucose supply, transport across the plasma membrane, and metabolism. To monitor glucose dynamics in individual cells, a glucose nanosensor was developed by flanking the Escherichia coli periplasmic glucose/galactose-binding protein with two different green fluorescent protein variants. Upon binding of substrate the FLIPglu-170n sensor showed a concentration-dependent decrease in fluorescence resonance energy transfer between the attached chromophores with a binding affinity for glucose of 170 nM. Fluorescence resonance energy transfer measurements with different sugars indicated a broad selectivity for monosaccharides. An affinity mutant with a K d of ϳ600 M was generated, which showed higher substrate specificity, and thus allowed specific monitoring of reversible glucose dynamics in COS-7 cells in the physiological range. At external glucose concentrations between 0.5 and 10 mM, reflecting typical blood levels, free cytosolic glucose concentrations remained at ϳ50% of external levels. The removal of glucose lead to reduced glucose levels in the cell, demonstrating reversibility and visualizing homeostasis. Glucose levels dropped even in the presence of the transport inhibitor cytochalasin B, indicating rapid metabolism. Consistently, the addition of 2-deoxyglucose, which is not recognized by the sensor, affects glucose uptake and metabolism rates. Within the physiological range, glucose utilization, i.e. hexokinase activity, was not limiting. Furthermore, the results show that in COS-7 cells, cytosolic glucose concentrations can vary over at least two orders of magnitude. The glucose nanosensor provides a novel tool with numerous scientific, medical, and environmental applications.

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Section: Fig 3 In Vivo Characterization Of Flipglu-600supporting

confidence: 56%

“…Pharmacological studies indicate that active transport via sodium/glucose cotransporter does not contribute significantly to glucose uptake (41). Hexokinase rather than glucokinase is used for glucose phosphorylation (32).…”

Section: Fig 5 Determining Cytosolic Glucose Concentrations At Diffmentioning

confidence: 99%

Imaging of the Dynamics of Glucose Uptake in the Cytosol of COS-7 Cells by Fluorescent Nanosensors

Fehr¹,

Lalonde

Lager

et al. 2003

Journal of Biological Chemistry

283

257

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“…Liver HXK4 occurs in a mitochondrial complex with WAVE-1, a multifunctional protein that regulates actin polymerization or that can be a protein kinase A anchoring protein (Danial et al, 2003). Liver glucokinase expressed in COS-7 cells was shown to colocalize with actin filaments (Murata et al, 1997). Also, HXK from bovine heart acts possibly as a capping protein in vitro when present during polymerization of G-actin (Wagner et al, 1999).…”

Section: Discussionmentioning

confidence: 99%

“…Notably, ACT2 and ACT8 proteins differ by only one amino acid. This was interesting because certain mammalian HXKs can bind to actin (Murata et al, 1997;Wagner et al, 1999). To test whether the corresponding Arabidopsis proteins can interact, we carried out coimmunoprecipitation assays between HXK1-FLAG and ACT2-HA ( Fig.…”

Section: Athxk1 Can Interact With Porin and Actin Proteinsmentioning

confidence: 99%

A Role for F-Actin in Hexokinase-Mediated Glucose Signaling

Balasubramanian

Karve²,

Kandasamy³

et al. 2007

Plant Physiology

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HEXOKINASE1 (HXK1) from Arabidopsis (Arabidopsis thaliana) has dual roles in glucose (Glc) signaling and in Glc phosphorylation. The cellular context, though, for HXK1 function in either process is not well understood. Here we have shown that within normal experimental detection limits, AtHXK1 is localized continuously to mitochondria. Two mitochondrial porin proteins were identified as capable of binding to overexpressed HXK1 protein, both in vivo and in vitro. We also found that AtHXK1 can be associated with its structural homolog, F-actin, based on their coimmunoprecipitation from transgenic plants that overexpress HXK1-FLAG or from transient expression assays, and based on their localization in leaf cells after cryofixation. This association might be functionally important because Glc signaling in protoplast transient expression assays is compromised by disruption of F-actin. We also demonstrate that Glc treatment of Arabidopsis seedlings rapidly and reversibly disrupts fine mesh actin filaments. The possible roles of actin in HXK-dependent Glc signaling are discussed.

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“…Analogous to other glycolytic enzymes, the translocation of GK to distinct subcellular compartments may be facilitated through the cytoskeleton. In line with this possibility, several glycolytic enzymes, including GK, have been shown to travel on actin filaments within cells (Murata et al, 1997;Giege et al, 2003;Waingeh et al, 2006;Balasubramanian et al, 2007). The actin cytoskeleton may help focus the glycolytic pathway toward sites of metabolic demand within the cell.…”

Section: Bad and Glucose Metabolismmentioning

confidence: 92%

BAD: undertaker by night, candyman by day

2008

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The BH3-only pro-apoptotic proteins are upstream sensors of cellular damage that selectively respond to specific, proximal death and survival signals. Genetic models and biochemical studies indicate that these molecules are latent killers until activated through transcriptional or post-translational mechanisms in a tissue-restricted and signal-specific manner. The large number of BH3-only proteins, their unique subcellular localization, protein-interaction network and diverse modes of activation suggest specialization of their damage-sensing function, ensuring that the core apoptotic machinery is poised to receive input from a wide range of cellular stress signals. The apoptotic response initiated by the activation of BH3-only proteins ultimately culminates in allosteric activation of pro-apoptotic BAX and BAK, the gateway proteins to the mitochondrial pathway of apoptosis. From activation of BH3-only proteins to oligomerization of BAX and BAK and mitochondrial outer membrane permeabilization, an intricate network of interactions between the pro-and anti-apoptotic members of the BCL-2 family orchestrates the decision to undergo apoptosis. Beyond regulation of apoptosis, multiple BCL-2 proteins have recently emerged as active components of select homeostatic pathways carrying other cellular functions. This review focuses on BAD, which was the first BH3-only protein linked to proximal survival signals through phosphorylation by survival kinases. In addition to findings that delineated the physiological role of BAD in apoptosis and its dynamic regulation by phosphorylation, studies pointing to new roles for this protein in other physiological pathways, such as glucose metabolism, are highlighted. By executing its 'day' and 'night' jobs in metabolism and apoptosis, respectively, BAD helps coordinate mitochondrial fuel metabolism and the apoptotic machinery.

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Co‐localization of glucokinase with actin filaments

Cited by 28 publications

References 19 publications

Imaging of the Dynamics of Glucose Uptake in the Cytosol of COS-7 Cells by Fluorescent Nanosensors

Imaging of the Dynamics of Glucose Uptake in the Cytosol of COS-7 Cells by Fluorescent Nanosensors

A Role for F-Actin in Hexokinase-Mediated Glucose Signaling

BAD: undertaker by night, candyman by day

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