Anaphase-promoting complex/cyclosome-Cdh1 coordinates glycolysis and glutaminolysis with transition to S phase in human T lymphocytes

Abstract

Cell proliferation is accompanied by an increase in the utilization of glucose and glutamine. The proliferative response is dependent on a decrease in the activity of the ubiquitin ligase anaphasepromoting complex/cyclosome (APC/C)-Cdh1 which controls G1-to-S-phase transition by targeting degradation motifs, notably the KEN box. This occurs not only in cell cycle proteins but also in the glycolysis-promoting enzyme 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase isoform 3 (PFKFB3), as we have recently demonstrated in cells in culture. We now show that APC/C-Cdh1 controls the proliferative response of human T lymphocytes. Moreover, we have found that glutaminase 1 is a substrate for this ubiquitin ligase and appears at the same time as PFKFB3 in proliferating T lymphocytes. Glutaminase 1 is the first enzyme in glutaminolysis, which converts glutamine to lactate, yielding intermediates for cell proliferation. Thus APC/C-Cdh1 is responsible for the provision not only of glucose but also of glutamine and, as such, accounts for the critical step that links the cell cycle with the metabolic substrates essential for its progression.cell cycle | glutaminase | 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase isoform 3 | proliferation H uman blood T lymphocytes have been used for many years in studies of cell proliferation (1, 2). These cells can be obtained directly from the circulation and therefore avoid the pitfalls associated with the use of cells in culture, which acquire confounding characteristics as a result of their in vitro environment (3). Interest has recently been rekindled in the metabolic changes that underpin cell proliferation in cancer to identify potential targets for chemotherapy (4,5). This has highlighted the need to carry out comparative studies on proliferating normal and tumor cells to ascertain whether an antimetabolic approach to cancer is possible without side effects related to the mechanism of action.The E3 ubiquitin ligase anaphase-promoting complex/cyclosome (APC/C) attached to the activator protein Cdh1 plays a crucial role in controlling G1-to S-phase transition, and therefore proliferation, through the breakdown of cell cycle proteins (6, 7). APC/C-Cdh1 substrates are targeted for degradation through specific recognition motifs, including one known as the KEN box (8). Inactivation of APC/C-Cdh1 in G1 of the cell cycle is necessary for initiation of S phase, in which DNA is replicated and chromosomes are duplicated. We have recently found that APC/CCdh1 links cell cycle activity with that of the enzyme 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase isoform 3 (PFKFB3) (9, 10). PFKFB3-a key regulator of glycolysis (11)-contains a KEN box motif and is thus also broken down by APC/C-Cdh1. Inactivation of APC/C-Cdh1 enables PFKFB3 to up-regulate glycolysis, thus providing the cell with the glucose essential for the subsequent biosynthesis of macromolecules. Our previous studies (9) were carried out in two cell lines; we therefore decided to investigate whether the same mechanis...

Two ubiquitin ligases, APC/C-Cdh1 and SKP1-CUL1-F (SCF)-β-TrCP, sequentially regulate glycolysis during the cell cycle

“…The function of this transient enhancement of glycolytic activity in late G1, which is necessary for cell cycle progression, now requires investigation because glycolysis might be used to generate energy, substrates for macromolecular synthesis, or for both purposes. Our present results obtained in a cancer-derived cell line, together with previous studies in a noncancer cell line (9) and in human T lymphocytes (10), suggest that these same, or similar, mechanisms underlie cell proliferation and enhanced metabolism in normal and neoplastic cells.…”

“…Antibodies used for immunoblotting included cyclin E from Novus, Cdh1 from Merck Chemicals, PFKFB3 from Abcam, β-TrCP from Cell Signaling Technology, PFK1 and cyclin A from Insight Biotechnology, and β-actin from Sigma. Generation of PFKFB3v5 antibodies has been described elsewhere (10).…”

“…We have recently found that PFKFB3 is regulated during cell proliferation by APC/C-Cdh1 and that silencing this enzyme prevents cells from entering S phase (9,10). These findings suggest that there is close coordination between cell cycle progression and provision of the raw materials required for its completion.…”

Uncoupling the Warburg effect from cancer

“…However, despite these observations, one might imagine that the Warburg effect need not be specific for cancer and that any normal cell would need to stimulate glycolysis to generate sufficient biosynthetic materials to fuel expansion and division. Recent work by Salvador Moncada's group published in PNAS (8) and other recent work from the same group (9, 10) provides exciting evidence supporting the idea that the Warburg effect is also required for the proliferation of noncancer cells.…”

“…1) (8). PFKFB3 potently stimulates glycolysis by catalyzing the formation of fructose-2,6-bisphosphate, the allosteric activator of 6-phosphofructo-1-kinase (11).…”