Disease relevance of Pklr

• These results strongly suggest that the mutant Pklr allele (Pklr(269A)) of AcB55/61 strains causes hemolytic anemia compensated by constitutive erythropoiesis, which in turn protects the mice against P. chabaudi infection [1].
• We used a recombinant adeno-associated virus (rAAV) that expresses a single-chain insulin analogue (SIA), which possesses biologically active insulin activity without enzymatic conversion, under the control of hepatocyte-specific L-type pyruvate kinase (LPK) promoter, which regulates SIA expression in response to blood glucose levels [2].
• To study the feasibility of gene therapy for PK deficiency, we first constructed the PK retrovirus pMNSM-hPK using human liver-type PK (LPK) cDNA and obtained a producer cell line of E86/AmPK [3].
• In this study, we demonstrated that xylitol at low concentration (O.5 mM) induced the expression of the L-PK/CAT construct in glucose-responsive mhAT3F hepatoma cells at the same level as 20 mM glucose, while it did not affect intracellular concentration of glucose 6-phosphate significantly [4].
• Pk1 adenylate cyclase readily elevated intracellular cAMP levels in mouse neuroblastoma cells (N1E-115) while Pk2EGTA adenylate cyclase had no effect on cAMP levels in these cells [5].

High impact information on Pklr

• Sequencing of candidate genes from the Char4 region identified a loss-of-function mutation (269T-->A, resulting in the amino acid substitution I90N) in the pyruvate kinase gene (Pklr) that underlies the malaria resistance in AcB55 and AcB61 [6].
• The aim of our study was to assess the role of ChREBP in the control of L-PK and FAS gene expression by PUFAs [7].
• Phosphorylation of the former resulted in inactivation of nuclear import, and that of the latter resulted in loss of the DNA-binding activity and L-PK transcription [8].
• Because RBC type PK is encoded by the Pk-1 locus of the mouse (chromosome 3), we designated the mutant locus as Pk-1slc [9].
• Northern blot analysis demonstrated the expression of the human LPK mRNA in each transduced cell line [3].

Biological context of Pklr

• Resistance in AcB55 and AcB61 is controlled by a locus on chromosome 3 (Char4) shown to be allelic with or tightly linked to a loss-of-function mutation in pyruvate kinase (Pklr) [1].
• Further studies using this method have identified a Pklr variant that confers resistance to murine malaria, a result that shows the potential of this approach to aid the understanding of mechanisms of disease resistance [10].
• The glucose-dependent activation of the L-PK gene is delayed, requires ongoing protein synthesis, and is mediated by the same glucose response element as in vivo and in isolated hepatocytes [11].
• Transfection of GLUT 2(-) cells with a GLUT 2 expression vector restored the inducibility of the L-PK promoter by glucose, mainly by suppressing the glucose-independent activity of this promoter [12].
• The functional role of the different sites binding transcriptional factors on the tissue-specific, glucose-responsive promoter of the L type pyruvate kinase gene (L-PK) has been investigated in transgenic mice [13].

Anatomical context of Pklr

• In addition, overexpression of a constitutive nuclear ChREBP isoform in cultured hepatocytes significantly reduced the PUFA inhibition of both L-PK and FAS gene expression [7].
• L-type pyruvate kinase (L-PK) is a key enzyme of the glycolytic pathway specifically expressed in the liver and, to a lesser degree, in the small intestine and kidney [14].
• The expression of human LPK enzyme activity was ascertained from the retrovirally transduced NIH/3T3 cells [3].
• The expression of the L-PK gene in GLUT 2(-) cells cultured in the absence of glucose was correlated with a high intracellular glucose 6-phosphate (Glu-6-P) concentration while under similar culture conditions Glu-6-P concentration was very low in GLUT 2(+) cells [12].
• These unprecedented results suggest that ChREBP rather than USF mediates glucose-promoted L-PK expression in insulin-secreting cells [15].

Associations of Pklr with chemical compounds

• It is concluded that Pk-1 is the structural gene for the erythrocyte and the major liver pyruvate kinase [16].
• ChREBP, essential for L-PK gene transcription, is activated by high glucose and inhibited by cAMP [8].
• The induction of L-PK caused an increase in the hepatic lactate concentration [17].
• Culture of GLUT 2(-) cells, in which the L-PK gene is constitutively expressed, in a culture medium using fructose as fuel selected GLUT 2(+) clones in which the L-PK gene responded to glucose [12].
• In hepatocytes in primary culture, 5 mM xylitol induced accumulation of the L-PK mRNA even in the absence of insulin [4].

Physical interactions of Pklr

• The liver/erythrocyte pyruvate kinase gene complex [Pk-1] in the mouse: regulatory gene mutations [18].

Analytical, diagnostic and therapeutic context of Pklr

• Electrophoretic mobility shift assays using the L-PK promoter segment reveal that induction of USF-1 and -2 dramatically enhances the USF binding activity, whereas DN-USF-1 and -2 abolish binding [15].
• Gel exclusion chromatography of Pk1 adenylate cyclase gave apparent Stokes radii (RS) of 43.5 A (+/- 1.3) in the presence of 2 mM CaCl2 and 33.8 A (+/- 0.94) in the presence of 2 mM EGTA [( ethylenebis (oxyethylenenitrilo)]tetraacetic acid) [5].
• METHODS: We established an immortalized cultured cortical TAL (cTAL) cell line from L-PK/Tag1 transgenic mice by microdissection [19].
• One of the MAb (154 IX B6) selected as a blood typing reagent agglutinated native P1 and Pk1 red cells with a high titer but was inactive against native P2, Pk2 and p erythrocytes [20].

References