Phosphotyrosine-containing lactate dehydrogenase is restricted to the nuclei of PC12 pheochromocytoma cells.

Abstract

There are five lactate dehydrogenase (LDH) isoenzymes, composed of various combinations of two types of subunits. LDH-5, which contains only the LDH A subunit, is known to be present in both the cytoplasm and the nucleus, to act as a single-stranded DNA-binding protein possibly functioning in transcription and/or replication, and to undergo phosphorylation of tyrosine 238 in approximately 1% of the enzyme after cell transformation by certain tumor viruses. We have characterized LDH from wild-type PC12 pheochromocytoma cells and from a PC12 variant (MPT1) that exhibits altered lactate metabolism and altered expression of multiple genes. Wild-type and MPT1 cells contain different proportions of LDH isoenzymes, with LDH-5 being more predominant in wild-type cells than in the variant. A small fraction of LDH from PC12 cells contains phosphotyrosine. Approximately 99% of the total LDH activity is located in the cytoplasm, but all of the phosphotyrosine-containing LDH is located in the nucleus. Furthermore, essentially all of the nuclear LDH contains phosphotyrosine. These results suggest that tyrosine phosphorylation can affect its role in the nucleus.Lactate dehydrogenase (LDH, EC 1.1.1.27) catalyzes the reversible conversion of pyruvate to lactate. Five LDH isoenzymes exist because the enzyme is a tetramer that is composed of various combinations of two kinds of Mr 35,000 subunits named A and B (9). The isoenzymes and their respective structures are LDH-5 (A4), LDH-4 (A3B), LDH-3 (A2B2), LDH-2 (AB3), and LDH-1 (B4). The LDH isoenzyme composition varies from tissue to tissue.In addition to its role in the metabolism of lactate and pyruvate, one of the LDH isoenzymes, LDH-5, may function in the regulation of gene transcription and/or DNA replication, since LDH-5 is present in the nucleus (5) and it has recently been found to be a previously studied singlestranded DNA-binding protein (14,25,31). It had been shown earlier that antibody raised against this DNA-binding protein cross-reacts with single-stranded DNA-binding proteins from heterologous species including Drosophila spp. (22). Furthermore, studies of Drosophila polytene salivary chromosomes revealed that this protein is associated nonrandomly with the chromosomes and is usually concentrated in certain chromosome puffs that were active sites of transcription, e.g., heat shock puffs after heat shock treatment (22). A possible role in replication is suggested by the finding that LDH-5 stimulates the activity of DNA polymerase a in vitro (14).Another LDH variation arises from phosphorylation. A small percentage of chicken fibroblast LDH-5 is phosphorylated at seine residues; after transformation of these cells by Rous sarcoma virus, 0.5 to 1% of LDH-5 is also phosphorylated at a single tyrosine residue, tyrosine 238 (6, 7). It is not clear that this tyrosine phosphorylation affects glycolysis, because such a small fraction of the LDH in the transformed cells contains phosphotyrosine. The intracellular location of phosphotyrosine-containing LDH-5 has not previously...

Tyrosine Phosphorylation of Lactate Dehydrogenase A Is Important for NADH/NAD+ Redox Homeostasis in Cancer Cells

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Increased tyrosine kinase activity of c-Src during calcium-induced keratinocyte differentiation.

“…Several recent reports suggest that tyrosine phosphorylation might regulate the function of some nuclear proteins. cdc2, lactose dehydrogenase, or androgen receptor are examples of nuclear proteins whose function or subcellular localization is affected by tyrosine phosphorylation (37)(38)(39). Furthermore, two other proteintyrosine kinases, the c-Abl IV protein and the FER gene product, have also been localized to the cell nucleus (40,41).…”

Evaluation of Myc E-Box Phylogenetic Footprints in Glycolytic Genes by Chromatin Immunoprecipitation Assays

“…Several isoforms of hexokinase play a key role in mitochondrion-mediated apoptosis by modulating proapoptotic molecules including Bax and Bad, suggesting that the glycolytic pathway and apoptosis are integrated (9,15,33). The roles of LDHA and GAPD proteins in glycolysis are well-established, yet the nuclear localization of these proteins suggest additional biological functions (45). Recently, GAPD and LDH were both found in a transcriptional coactivator complex that assists the Oct-1 transcription factor in regulating histone H2B expression (44).…”