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  • Glucose metabolic pathways are well defined but the subcellu

    2021-11-30

    Glucose metabolic pathways are well defined but the subcellular organization of these pathways is poorly understood in liver and kidney. Several factors affect the net flux of gluconeogenesis and allow the recently to respond to physiological changes [1]. Ovadi and Srere have suggested that a new regulatory mechanism for metabolic pathways is the subcellular compartmentalization and the physical separation of them in a single cell [17], [18]. These authors also underline the need for a thorough characterization of the subcellular organization of metabolic enzymes [17], [18]. In this context, in muscle there is convincing evidence about the interaction between some glycolytic enzymes and the subcellular cytoskeletal network, association that regulates the location of glycolytic enzymes and the kinetic rates of individual enzymes in the contraction process [19], [20], [21]. In this report, we present the first data available on the subcellular localization of liver FBPase in rat kidney and liver cells that clearly describe the localization of FBPase in the nucleus and in a compartment near the apical membrane in kidney cells and in an area adjacent to the plasma membrane in the hepatocytes. Which are the physiological reasons for the localization of FBPase in the periphery of these cells? Kidney contributes with a significant fraction of the systemic gluconeogenesis [22], [23]. Recently, we have established that liver FBPase and C-PEPCK are expressed mainly in the human proximal tubule cells localized in the kidney cortex, indicating that the glucose synthesis is compartmentalized to the proximal tubules [10], [11]. A similar distribution of glycolytic and gluconeogenic enzymes has been described in kidney and liver [11], [12], [24], [25], suggesting that the separate localization of both pathways within the nephron, coupled with the elevation of renal precursors in the proximal tubule, may represent an additional physiological mechanism that stimulates the gluconeogenic pathway and avoids the consumption of ATP by the formation of futile cycles in a single cell [10], [11]. Moreover, this analysis suggests that the presence of these catalytically active key enzymes is not sufficient to explain the mechanism responsible for the variation in gluconeogenic rates from lactate in the proximal tubules. Our findings [10], showing the expression of the monocarboxylated transporter 1 in the S1 segment, may explain these different rates in the proximal tubule segments. The FBPase shows a specialized subcellular compartmentalization along rat kidney proximal tubules, being clearly localized in the apical area of these cells. The treatment of a kidney crude extract with Triton X-100 led to a significant increase of 250% of the total activity of FBPase. From these data we propose that FBPase may interact with transport elements in the cellular membrane and play an important function in the metabolic heterogeneity observed among the proximal tubules. This functional duality of gluconeogenic enzymes has been demonstrated for aldolase [26], [27]. Indeed, this enzyme plays a structural role in the polymerization of actin and in the dynamic association of GLUT 4 vesicles with actin, regulating glucose transport [26], [27]. Also, we can postulate that glucose synthesis in these cells occurs in a subcellular compartment adjacent to the apical membrane. A similar metabolic zonation has been observed in rat liver, demonstrating that hepatocytes from the periportal region of rat liver mainly expressed the gluconeogenic enzymes, whereas the hepatocytes located in the pericentral region mainly expressed glycolytic enzymes [11], [24], [28], [29]. These data and other interesting results strengthen the notion of the importance of different metabolic functions for particular cells in bi-functional organs [10], [11], [24], [30]. The negative immunostaining in non-hepatocyte cells is a good control for our antiserum and supports the specificity of our subcellular localization. Interestingly, FBPase was located at the periphery of the plasma membrane of adjacent hepatocytes. Studies by Guinovart’s laboratory have demonstrated that liver glycogen synthase (GS) has a cytosolic distribution in the absence of glucose and concentrates at the periphery of the hepatocyte when the concentration of the hexose increases [31]. Additionally, these authors showed that GS colocalizes with the glycogen deposit stores in the membrane compartment, suggesting that this enzyme remains attached to its product [32]. Glucose depletion causes glycogen degradation and the redistribution of the GS to the cytosol [33]. Our results show that the intracellular distribution of FBPase closely resembles the GS and glycogen localization and lead us to propose that FBPase in this compartment is participating in the production of glucose 6-phosphate as a precursor for glycogen synthesis. Therefore, the existence of this organized subcellular distribution and the possible separation recently of glycogenogenesis and gluconeogenesis pathways within a single cell might be the regulatory mechanism that may explain how the kidney and hepatic cells can segregate intermediates of competing metabolic pathways. This idea is sustained by the capacity of the liver GS to differentiate between glucose 6-phosphate produced by glucokinase (GK) or hexokinase I and by the capacity of GS to use the glucose 6-phosphate produced by gluconeogenesis from dihydroxyacetone [34]. Together these observations support the proposal that the physical separation of FBPase in multiple compartments and its interaction with enzymes that participate in glycogen synthesis and gluconeogenesis are required for the regulation of these pathways.