Autophagy provides nutrients but can lead to Chop-dependent induction of Bim to sensitize growth factor-deprived cells to apoptosis.

Tissue homeostasis is controlled by the availability of growth factors, which sustain exogenous nutrient uptake and prevent apoptosis. Although autophagy can provide an alternate intracellular nutrient source to support essential basal metabolism of apoptosis-resistant growth factor-withdrawn cells, antiapoptotic Bcl-2 family proteins can suppress autophagy in some settings. Thus, the role of autophagy and interactions between autophagy and apoptosis in growth factor-withdrawn cells expressing Bcl-2 or Bcl-xL were unclear. Here we show autophagy was rapidly induced in hematopoietic cells upon growth factor withdrawal regardless of Bcl-2 or Bcl-xL expression and led to increased mitochondrial lipid oxidation. Deficiency in autophagy-essential gene expression, however, did not lead to metabolic catastrophe and rapid death of growth factor-deprived cells. Rather, inhibition of autophagy enhanced survival of cells with moderate Bcl-2 expression for greater than 1 wk, indicating that autophagy promoted cell death in this time frame. Cell death was not autophagic, but apoptotic, and relied on Chop-dependent induction of the proapoptotic Bcl-2 family protein Bim. Therefore, although ultimately important, autophagy-derived nutrients appear initially nonessential after growth factor withdrawal. Instead, autophagy promotes tissue homeostasis by sensitizing cells to apoptosis to ensure only the most apoptosis-resistant cells survive long-term using autophagy-derived nutrients when growth factor deprived.

Glucose metabolism attenuates p53 and Puma-dependent cell death upon growth factor deprivation.

Growth factor stimulation and oncogenic transformation lead to increased glucose metabolism that may provide resistance to cell death. We have previously demonstrated that elevated glucose metabolism characteristic of stimulated or cancerous cells can stabilize the anti-apoptotic Bcl-2 family protein Mcl-1 through inhibition of GSK-3. Here we show that the pro-apoptotic Bcl-2 family protein, Puma, is also metabolically regulated. Growth factor deprivation led to the loss of glucose uptake and induction of Puma. Maintenance of glucose uptake after growth factor withdrawal by expression of the glucose transporter, Glut1, however, suppressed Puma up-regulation and attenuated growth factor withdrawal-induced activation of Bax, DNA fragmentation, and cell death. Conversely, glucose deprivation led to Puma induction even in the presence of growth factor. This regulation of Puma expression was a central component in cell death as a consequence of growth factor or glucose deprivation because Puma deficiency suppressed both of these cell death pathways. Puma induction in growth factor or glucose withdrawal was dependent on p53 in cell lines and in activated primary T lymphocytes because p53 deficiency suppressed Puma induction and delayed Bax and caspase activation, DNA fragmentation, and loss of clonogenic survival. Importantly, although p53 levels did not change or were slightly reduced, p53 activity was suppressed by elevated glucose metabolism to inhibit Puma induction after growth factor withdrawal. These data show that p53 is metabolically regulated and that glucose metabolism initiates a signaling mechanism to inhibit p53 activation and suppress Puma induction, thus promoting an anti-apoptotic balance to Bcl-2 family protein expression that supports cell survival.

New roles for pyruvate kinase M2: working out the Warburg effect.

The origins and role of the Warburg effect have remained uncertain for many years. Two recent studies demonstrate that an embryonic- and cancer-cell-specific isoform of the enzyme pyruvate kinase M2 (PKM2) is regulated by binding to phospho-tyrosine motifs and promotes increased cell growth and tumor development. PKM2 enhances the use of glycolytic intermediates for macromolecular biosynthesis and tumor growth. These findings illustrate the distinct advantages of this metabolic phenotype in cancer cell growth.

Activation of Wnt signaling in hematopoietic regeneration.

Hematopoietic stem cells (HSCs) respond to injury by rapidly proliferating and regenerating the hematopoietic system. Little is known about the intracellular programs that are activated within HSCs during this regenerative process and how this response may be influenced by alterations in signals from the injured microenvironment. Here we have examined the regenerating microenvironment and find that following injury it has an enhanced ability to support HSCs. During this regenerative phase, both hematopoietic and stromal cell elements within the bone marrow microenvironment show increased expression of Wnt10b, which can function to enhance growth of hematopoietic precursors. In addition, regenerating HSCs show increased activation of Wnt signaling, suggesting that microenvironmental changes in Wnt expression after injury may be integrated with the responses of the hematopoietic progenitors. Cumulatively, our data reveal that growth signals in the hematopoietic system are re-activated during injury, and provide novel insight into the influence of the microenvironment during regeneration.

Effect of ertapenem protein binding on killing of bacteria.

The effect of protein binding on the antimicrobial activity of ertapenem was evaluated using the bacterial kill rate and concentration-response studies. Various proportions of human serum were utilized to determine the total and free-drug concentrations using a validated high-performance liquid chromatography assay. The MICs and kill curves were determined for test isolates of Enterobacter cloacae and Staphylococcus aureus at various percentages of human serum. The killing of bacteria was analyzed in relation to the free and total concentrations of ertapenem at various proportions of human serum. It was determined that unbound ertapenem was responsible for the antimicrobial activity against the test isolates.