Molecular cloning and expression of rat aldolase C messenger RNA during development and hepatocarcinogenesis

Skala, Henriette; Vibert, M.; Lamas, Eugenia; Maire, Pascal; Schweighoffer, Fabien; Kahn, Axel

doi:10.1111/j.1432-1033.1987.tb10898.x

Cited by 26 publications

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References 34 publications

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“…The 3.2-kb fragment hybridized (Fig. l a ) with r.Al.C.2 cDNA which contains the 3' noncoding region of the mRNA [13]. The 2.2-kb fragment hybridized at low stringency with an aldolase B cDNA clone, previously isolated in our laboratory (Besmond, C., unpublished results), which covers the 5' coding extremity of the messenger, coded by exon 11.…”

Section: Isolation and Characterization Of The Rat Aldolase C Genementioning

confidence: 99%

“…Approximately 1.2 x 1 O6 recombinant bacteriophages were screened with the nick-translated, previously isolated probe r.Al.C.l [13]. This probe covers 340 bases in the 3' coding part of the mRNA.…”

Section: Isolation and Characterization Of The Rat Aldolase C Genementioning

confidence: 99%

“…Escherichia coli strain EC 803 was infected by the phages and screened by in sifu plaque hybridization [21] with the nick-translated r.AI.C.l cDNA probe [13]. One clone containing a 15-kb insert was isolated and purified on two successive cesium chloride gradients.…”

Section: Screening Of a Rat Genomic Librarymentioning

confidence: 99%

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The brain‐specific gene for rat aldolase C possesses an unusual housekeeping‐type promoter

Vibert¹,

Henry²,

Kahn³

et al. 1989

European Journal of Biochemistry

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A DNA fragment encompassing the first exon and about 750 bp of the 5'-flanking sequence has been isolated and sequenced. The gene has multiple start sites of transcription which are dispersed over about 200 bp. The promoter lacks TATA and CAAT boxes and is very G + C-rich, with putative binding sites for the transcriptional factors Spl and AP2. Similar features are shared with two other brain-specific genes encoding thy-1 antigen and y-enolase. The existence of a conserved block of similarity upstream of the human and rat aldolase C genes suggests that this region could be involved in tissue-specific expression whose mechanism seem to be, at least in part, transcriptional.Fructose 1,6-bisphosphate aldolase, a glycolytic enzyme, exists as three types: aldolase A (muscel type), aldolase B (liver type) and aldolase C (brain type). Expression of the isozymes in various tissues, and changes of the isozymic pattern during development and carcinogenesis, have been extensively studied in several species [I -41. Aldolase C is strongly expressed in aduit and foetal brain, and, at a lower level, in all foetal tissues and in some pathological situations such as chemically induced rat hepatocarcinoma [4].It is clear that all the aldolase genes are derived by duplication from a common ancestral gene, the coding sequences between the different members of this family being at least 70% similar and the exon-intron structures being conserved. The tissue specificities of these genes are therefore expected to result from different control regions, acting mainly at the transcriptional level. We have consequently begun to investigate systematically the structure and function of the 5'-flanking region of the different aldolase genes.Previously, we and others have isolated complementary DNA clones for liver aldolase B [5 -81, muscle aldolase A [9, 101 and brain aldolase C [ll-131. Genes for the three aldolases have been cloned from different species and their structure has been analyzed [15 -201. In the present work, we report the isolation and characterization of the rat aldolase C gene. The promoter region of the gene was sequenced and the start sites of transcription were mapped on RNAs from different tissues.The structure of the 5'-flanking region proved to be very unusual for a tissue-specific gene, with highly heterogeneous start sites of transcription and absence of TATA and CAAT boxes. Nevertheless, the molecular basis of the tissue-specific expression of this gene seems to be, at least in part, transcriptional. Correspondence to MATERIALS AND METHODS Screening of a rat genomic libraryA genomic DNA library containing an HaeIII partial digest of rat DNA A charon 4A was kindly provided by Linda Jago-Sinlki. Escherichia coli strain EC 803 was infected by the phages and screened by in sifu plaque hybridization [21] with the nick-translated r.AI.C.l cDNA probe [13]. One clone containing a 15-kb insert was isolated and purified on two successive cesium chloride gradients. DNA was extracted with phenol after proteinase K digestion. R...

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Section: Isolation and Characterization Of The Rat Aldolase C Genementioning

confidence: 99%

Section: Isolation and Characterization Of The Rat Aldolase C Genementioning

confidence: 99%

See 1 more Smart Citation

The brain‐specific gene for rat aldolase C possesses an unusual housekeeping‐type promoter

Vibert¹,

Henry²,

Kahn³

et al. 1989

European Journal of Biochemistry

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“…Data from former reports suggest that there are many factors contributing to the origin of aerobic glycolysis (2). The altered control of glycolysis by expression of certain isoenzymes is one important factor (2)(3)(4)(5)(6)(7)(8)(9)(10)(11)(12). Furthermore, the glycerol 3-phosphate shuttle and the malate-aspartate shuttle are altered in such a way that transport of cytosolic hydrogen into the mitochondria is reduced, requiring tumor cells to reoxidize NADH cytosolically by lactate dehydrogenase (13)(14)(15).…”

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Effect of Extracellular AMP on Cell Proliferation and Metabolism of Breast Cancer Cell Lines with High and Low Glycolytic Rates

Mazurek

Michel²,

Eigenbrodt

1997

Journal of Biological Chemistry

101

107

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In differentiated tissues, such as muscle and brain, increased adenosine monophosphate (AMP) levels stimulate glycolytic flux rates. In the breast cancer cell line MCF-7, which characteristically has a constantly high glycolytic flux rate, AMP induces a strong inhibition of glycolysis. The human breast cancer cell line MDA-MB-453, on the other hand, is characterized by a more differentiated metabolic phenotype. MDA-MB-453 cells have a lower glycolytic flux rate and higher pyruvate consumption than MCF-7 cells. In addition, they have an active glycerol 3-phosphate shuttle. AMP inhibits cell proliferation as well as NAD and NADH synthesis in both MCF-7 and MDA-MB-453 cells. However, in MDA-MB-453 cells glycolysis is slightly activated by AMP. This disparate response of glycolytic flux rate to AMP treatment is presumably caused by the fact that the reduced NAD and NADH levels in AMP-treated MDA-MB-453 cells reduce lactate dehydrogenase but not cytosolic glycerol-3-phosphate dehydrogenase reaction. Due to the different enzymatic complement in MCF-7 cells, proliferation is inhibited under glucose starvation, whereas MDA-MB-453 cells grow under these conditions. The inhibition of cell proliferation correlates with a reduction in glycolytic carbon flow to synthetic processes and a decrease in phosphotyrosine content of several proteins in both cell lines.Both proliferating cells and tumor cells maintain a high glycolytic rate even under aerobic conditions, a process referred to as aerobic glycolysis. Observations on aerobic glycolysis in tumor cells prompted Warburg (1) to postulate an altered respiratory function leading to an increased glycolytic capacity and a high rate of lactate formation from glucose in the presence of oxygen. Data from former reports suggest that there are many factors contributing to the origin of aerobic glycolysis (2). The altered control of glycolysis by expression of certain isoenzymes is one important factor (2-12). Furthermore, the glycerol 3-phosphate shuttle and the malate-aspartate shuttle are altered in such a way that transport of cytosolic hydrogen into the mitochondria is reduced, requiring tumor cells to reoxidize NADH cytosolically by lactate dehydrogenase (13-15). Additionally, oxidation of pyruvate is reduced in favor of glutamine oxidation (16 -25). Due to the expression of the mitochondrial, NAD-dependent malate decarboxylase, malate is converted to pyruvate and lactate (22)(23)(24). The conversion of glutamine to lactate is called, in analogy to glycolysis, glutaminolysis (25). In tumor cells the glycolytic capacity can be so great that all of the cell's energy requirements are derived from glycolysis (2, 26). Therefore, high glycolytic activity ensures the survival and the migration of tumor cells in hypoxic areas (2,26,27). The main role of the glutaminolytic pathway is the generation of energy (2, 25). However, a high glycolytic rate is not always linked to cell proliferation or tumor formation. There are several cell lines that are able to grow in a medium with 5 mM gala...

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“…These closely related isoenzymes, designated aldolase A, B and C, are expressed in a tissue-specific fashion and are regulated during ontogenesis and cancerogenesis [2,3]. Three genes, A, B and C, locaiized on chromosomes 16% 9 and 17, respectively, encode for the three ~so~~z~mes (ih,S,@ We and others have eIoned and sequenced the human genes and cDNAs for the three aldolases [7--l 7].…”

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Cis‐acting elements in the promoter region of the human aldolase C gene

et al. 1993

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We i~vesti~f~d the c&acting sequences involve m the expression of the human Adolph C gene by transient tmnsf~t~ons into b~man neuroblastoma cells (SKNBE). We demonstrate that 420 bp of the Y-flanking DNA dimct at high efficiency the transcription of the CAT reporter gene. A deletion between -420 bp and -164 bp causes a 60% decrease ofCAT activity. Gel shift and DNase I footprmtmg analyses revealed four protected elements: A, B, C and D. Competition analyses indicate that Spl or factors sharing a similar sequence specificity bind to elements A and B, but not to elements C and D. Sequence analysts shows a half palindromic ERE motrf(GGTCA), in elements B and D. Region D binds a transactivatmg factor which appears also essential to stabtlize the initiation complex.Aldolase C; Gene expression; Neuroblastoma cell (SKNBE), &-acting elements; Spl transacting factor Three distinct types of mammalian fructose-1 ,6-bisphosphate aldolase have been characterized by a variety of structural, biochemical and immunological methods [l]. These closely related isoenzymes, designated aldolase A, B and C, are expressed in a tissue-specific fashion and are regulated during ontogenesis and cancerogenesis [2,3]. Three genes, A, B and C, locaiized on chromosomes 16% 9 and 17, respectively, encode for the three ~so~~z~mes (ih,S,@ We and others have eIoned and sequenced the human genes and cDNAs for the three aldolases [7--l 7].Aldolase C is eharaGteristic of brain, where it is mainly localized in neurons and astroghal cells fl8.19]. The protein sequence of aldolase C has been deduced from the nucleotide sequence of mouse and rat complementary cDNA clones [20,21] and from human and rat genomic clones [13,14,17,22]. The 5' non-coding region has been analyzed in rat and man. Multiple start sites of transcription have been reported in rat f22], and one major start site has been described in man ]17].In this paper we report the analysis of the ~~ornot~r region performed by ~~a~s~~~~* of fusion constructs with a reporter gene into human neuronaf (SKNBE) cefl line and the identification of the bus-elements involved in the regulation of the human aldoias~ C gene expression MATERIALS AND METHODS I.. Phwtid constructionThe aldolase C promoter region was excused from the gene as an XbaIlPsfI fragment. spanning from -1190 bp to +20 relative to the major transc~pt~onaI start site fI7], and cloned into the Ml3 rap19 vector. The deletions were made by cuttmg the Y-end with X&rI and subsequently digesting with ExQIII m&ease (Boehringer Ma~n~e~m) @3]. The digested fragments were flush-ended by Klenow poiymerase and hgated into the Mi3 mp19 vector. The deleted fragments were checked by sequencing wtth the chain te~~nat~on method 124], excised from the vector with the appropriate res~~~ct~on enzyme, and then cloned m SmaIlPsrI sites upstream from the CAT reporter gene in the pEMBL-&CAT expression vector [25]. Chrmeric aldolase C-CAT plasmids were purified on a cesium chloride gradient. Cell culture, truxrrent trunsfections and CAT assaysHuman neuroblastoma ...

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Molecular cloning and expression of rat aldolase C messenger RNA during development and hepatocarcinogenesis

Cited by 26 publications

References 34 publications

The brain‐specific gene for rat aldolase C possesses an unusual housekeeping‐type promoter

The brain‐specific gene for rat aldolase C possesses an unusual housekeeping‐type promoter

Effect of Extracellular AMP on Cell Proliferation and Metabolism of Breast Cancer Cell Lines with High and Low Glycolytic Rates

Cis‐acting elements in the promoter region of the human aldolase C gene

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