Overflow metabolism refers to the seemingly wasteful strategy in which cells use fermentation instead of the more efficient respiration to generate energy, despite the availability of oxygen. Known as Warburg effect in the context of cancer growth, this phenomenon occurs ubiquitously for fast growing cells, including bacteria, fungi, and mammalian cells, but its origin has remained mysterious despite decades of research. Here we study metabolic overflow in E. coli and show that it is a global physiological response used to cope with changing proteomic demands of energy biogenesis and biomass synthesis under different growth conditions. A simple model of proteomic resource allocation can quantitatively account for all of the observed behaviors and accurately predict responses to novel perturbations. The key hypothesis of the model, that the proteome cost of energy biogenesis by respiration exceeds that by fermentation, is quantitatively confirmed by direct measurement of protein abundances via quantitative mass spectrometry.

Summary. Background: Platelet glycoprotein (GP)VI that binds collagen, and GPIb-IX-V that binds von Willebrand factor, initiate thrombus formation. Objectives: In this study, we investigated the mechanisms of metalloproteinase-mediated ectodomain shedding that regulate the surface expression of GPVI, GPIba (the major ligand-binding subunit) and GPV (that regulates thrombin-dependent activation via GPIba). Methods and results: Immunoblotting human platelet lysates using affinity-purified antibodies against cytoplasmic domains of GPVI, GPIba or GPV allowed simultaneous analysis of intact and cleaved receptor, and revealed (i) that a significant fraction of GPIba, but not GPVI, exists in a cleaved state on platelets, even when isolated in the presence of metalloproteinase inhibitor (GM6001) or EDTA; (ii) the samesized membrane-associated fragments of GPVI or GPIba are generated by phorbol-ester (PMA), the mitochondrial-targeting reagent CCCP, the calmodulin inhibitor W7, or the thiolmodifying reagent, N-ethylmaleimide, that directly activates ADAM10/ADAM17; and (iii) GPV is shed by both metalloproteinase-and thrombin-dependent mechanisms, depending on the concentration of thrombin. Based on the predicted cleavage area defined by these studies, ADAM10, but not ADAM17, cleaved a GPVI-based synthetic peptide within the extracellular membrane-proximal sequence (PAR^Q

No abstract

The glycoprotein VI (GPVI)-Fc receptor (FcR)␥The adhesion and activation of platelets by subendothelial collagen fibers initiates aggregate formation at sites of vessel damage. Glycoprotein (GP) 1 VI plays a critical role in the activatory events induced by collagen as shown by the lack of response to collagen in human and mice platelets deficient in the glycoprotein (1, 2). A collagen-related peptide and a snake venom toxin, convulxin, interact specifically with GPVI and mimic many of the responses to collagen (3-5).Because of the physiological importance of GPVI, the mechanism of the GPVI-mediated signaling system has been extensively investigated (6 -8). GPVI is present as a complex with Fc receptor (FcR) ␥-chain in the platelet membrane (8 -10). The Src family kinases, Fyn and Lyn, are associated with GPVIFcR ␥-chain complex in platelets and initiate activation through phosphorylation of the immunoreceptor tyrosinebased activation motif (ITAM) in the FcR ␥-chain leading to binding and activation of the tyrosine kinase Syk. A series of adapter molecules including LAT and SLP76 orchestrate a carefully regulated signaling network leading to activation of PLC␥2, phosphoinositol 3-kinase, and small molecular weight G proteins, leading to platelet activation (6, 7).The cloning of GPVI (11-14) has revealed it to be a member of the immunoglobulin (Ig) superfamily, showing close homology to Fc␣RI. GPVI has a charged arginine residue in its transmembrane domain. This arginine, together with elements within the cytoplasmic domain, is crucial for association of GPVI with FcR ␥-chain and GPVI-mediated signal transduction (15,16). In addition, the cytoplasmic tail of GPVI has a cluster of 6 proline residues of unknown function (11)(12)(13)(14). This sequence of GPVI, RPLPPLPPLP, contains a consensus Src family kinase-SH3 recognition motif (RPLPPLP) (17,18), and provides a potential site of interaction with Fyn and Lyn via their SH3 domains.In this study, we demonstrate that depletion of the prolinerich domain in GPVI abolishes the association with Fyn and Lyn and prevents tyrosine phosphorylation of FcR ␥-chain and downstream responses. From these findings, we suggest that Fyn/Lyn directly bind the proline-rich domain of GPVI and that this association is necessary for phosphorylation of the ITAM and downstream signals.

We present the analysis of the evolution of tumors in a case of hepatocellular carcinoma. This case is particularly informative about cancer growth dynamics and the underlying driving mutations. We sampled nine different sections from three tumors and seven more sections from the adjacent nontumor tissues. Selected sections were subjected to exon as well as whole-genome sequencing. Putative somatic mutations were then individually validated across all 9 tumor and 7 nontumor sections. Among the mutations validated, 24 were amino acid changes; in addition, 22 large indels/copy number variants (>1 Mb) were detected. These somatic mutations define four evolutionary lineages among tumor cells. Separate evolution and expansion of these lineages were recent and rapid, each apparently having only one lineage-specific protein-coding mutation. Hence, by using a cell-population genetic definition, this approach identified three coding changes (CCNG1, P62, and an indel/fusion gene) as tumor driver mutations. These three mutations, affecting cell cycle control and apoptosis, are functionally distinct from mutations that accumulated earlier, many of which are involved in inflammation/immunity or cell anchoring. These distinct functions of mutations at different stages may reflect the genetic interactions underlying tumor growth.cell genealogy | cellular evolution | foreground mutation T umorigenesis is generally believed to be the consequence of mutation accumulation, including single nucleotide substitutions, structural variations, and epigenetic changes, in somatic cells (1). A typical cancer may have thousands of somatic mutations, of which 10-100 may be in coding regions (2-7). A central issue in cancer genomics is then the dynamics of tumor growth in relation to the accumulation of these mutations. Given any individual case of cancer, the questions are hence: (i) how many adaptive mutations drive the tumor growth; (ii) how strongly each mutation drives the growth; and (iii) what their molecular nature is vis-à-vis that of the background mutations. To answer these questions, we treat each tumor as a population of cells and apply population genetic principles to infer adaptive mutations (8).Cancer mutations are often divided into drivers and passengers (9). Driver mutations are those that contribute directly to tumorigenesis and their identification is crucial for understanding the molecular biology of cancers. An important issue is how driver mutations should be defined operationally. Candidate driver mutation in the literature often refers to coding changes in genes that are commonly mutated, for example, in multiple cases of hepatocellular carcinoma (HCC). Adaptive mutation proposed here is an alternative definition of candidate driver mutation, inferred from the dynamics of cell proliferation in its natural setting within a single patient.In this report, we analyze a case of HCC, the fifth most common cancer worldwide, by such an approach. We regard HCC as particularly favorable for identifying candidate driver mutatio...