Pathogenicity and virulence of Listeria monocytogenes: A trip from environmental to medical microbiology.

Listeria monocytogenes is a saprophytic gram-positive bacterium, and an opportunistic foodborne pathogen that can produce listeriosis in humans and animals. It has evolved an exceptional ability to adapt to stress conditions encountered in different environments, resulting in a ubiquitous distribution. Because some food preservation methods and disinfection protocols in food-processing environments cannot efficiently prevent contaminations, L. monocytogenes constitutes a threat to human health and a challenge to food safety. In the host, Listeria colonizes the gastrointestinal tract, crosses the intestinal barrier, and disseminates through the blood to target organs. In immunocompromised individuals, the elderly, and pregnant women, the pathogen can cross the blood-brain and placental barriers, leading to neurolisteriosis and materno-fetal listeriosis. Molecular and cell biology studies of infection have proven L. monocytogenes to be a versatile pathogen that deploys unique strategies to invade different cell types, survive and move inside the eukaryotic host cell, and spread from cell to cell. Here, we present the multifaceted Listeria life cycle from a comprehensive perspective. We discuss genetic features of pathogenic Listeria species, analyze factors involved in food contamination, and review bacterial strategies to tolerate stresses encountered both during food processing and along the host's gastrointestinal tract. Then we dissect host-pathogen interactions underlying listerial pathogenesis in mammals from a cell biology and systemic point of view. Finally, we summarize the epidemiology, pathophysiology, and clinical features of listeriosis in humans and animals. This work aims to gather information from different fields crucial for a comprehensive understanding of the pathogenesis of L. monocytogenes.

Assessment of the interaction between the flux-signaling metabolite fructose-1,6-bisphosphate and the bacterial transcription factors CggR and Cra.

Bacteria regulate cell physiology in response to extra- and intracellular cues. Recent work showed that metabolic fluxes are reported by specific metabolites, whose concentrations correlate with flux through the respective metabolic pathway. An example of a flux-signaling metabolite is fructose-1,6-bisphosphate (FBP). In turn, FBP was proposed to allosterically regulate master regulators of carbon metabolism, Cra in Escherichia coli and CggR in Bacillus subtilis. However, a number of questions on the FBP-mediated regulation of these transcription factors is still open. Here, using thermal shift assays and microscale thermophoresis we demonstrate that FBP does not bind Cra, even at millimolar physiological concentration, and with electrophoretic mobility shift assays we also did not find FBP-mediated impairment of Cra's affinity for its operator site, while fructose-1-phosphate does. Furthermore, we show for the first time that FBP binds CggR within the millimolar physiological concentration range of the metabolite, and decreases DNA-binding activity of this transcription factor. Molecular docking experiments only identified a single FBP binding site CggR. Our results provide the long thought after clarity with regards to regulation of Cra activity in E. coli and reveals that E. coli and B. subtilis use distinct cellular mechanism to transduce glycolytic flux signals into transcriptional regulation.

Metabolic-flux dependent regulation of microbial physiology.

According to the most prevalent notion, changes in cellular physiology primarily occur in response to altered environmental conditions. Yet, recent studies have shown that changes in metabolic fluxes can also trigger phenotypic changes even when environmental conditions are unchanged. This suggests that cells have mechanisms in place to assess the magnitude of metabolic fluxes, that is, the rate of metabolic reactions, and use this information to regulate their physiology. In this review, we describe recent evidence for metabolic flux-sensing and flux-dependent regulation. Furthermore, we discuss how such sensing and regulation can be mechanistically achieved and present a set of new candidates for flux-signaling metabolites. Similar to metabolic-flux sensing, we argue that cells can also sense protein translation flux. Finally, we elaborate on the advantages that flux-based regulation can confer to cells.

Non-coding RNA regulation in pathogenic bacteria located inside eukaryotic cells.

Intracellular bacterial pathogens have evolved distinct lifestyles inside eukaryotic cells. Some pathogens coexist with the infected cell in an obligate intracellular state, whereas others transit between the extracellular and intracellular environment. Adaptation to these intracellular lifestyles is regulated in both space and time. Non-coding small RNAs (sRNAs) are post-transcriptional regulatory molecules that fine-tune important processes in bacterial physiology including cell envelope architecture, intermediate metabolism, bacterial communication, biofilm formation, and virulence. Recent studies have shown production of defined sRNA species by intracellular bacteria located inside eukaryotic cells. The molecules targeted by these sRNAs and their expression dynamics along the intracellular infection cycle remain, however, poorly characterized. Technical difficulties linked to the isolation of "intact" intracellular bacteria from infected host cells might explain why sRNA regulation in these specialized pathogens is still a largely unexplored field. Transition from the extracellular to the intracellular lifestyle provides an ideal scenario in which regulatory sRNAs are intended to participate; so much work must be done in this direction. This review focuses on sRNAs expressed by intracellular bacterial pathogens during the infection of eukaryotic cells, strategies used with these pathogens to identify sRNAs required for virulence, and the experimental technical challenges associated to this type of studies. We also discuss varied techniques for their potential application to study RNA regulation in intracellular bacterial infections.

The Listeria Small RNA Rli27 Regulates a Cell Wall Protein inside Eukaryotic Cells by Targeting a Long 5'-UTR Variant.

Listeria monocytogenes is a bacterial pathogen whose genome encodes many cell wall proteins that bind covalently to peptidoglycan. Some members of this protein family have a key role in virulence, and recent studies show that some of these, such as Lmo0514, are upregulated in bacteria that colonize eukaryotic cells. The regulatory mechanisms that lead to these changes in cell wall proteins remain poorly characterized. Here we studied the regulation responsible for increased Lmo0514 protein levels in intracellular bacteria. The amount of this protein increased markedly in intracellular bacteria (>200-fold), which greatly exceeded the increase in lmo0514 transcript levels (∼6-fold). Rapid amplification of 5'-cDNA ends (RACE) assays identified two lmo0514 transcripts with 5'-untranslated regions (5'-UTR) of 28 and 234 nucleotides. The transcript containing the long 5'-UTR is upregulated by intracellular bacteria. The 234-nucleotide 5'-UTR is also the target of a small RNA (sRNA) denoted Rli27, which we identified by bioinformatics analysis as having extensive base pairing potential with the long 5'-UTR. The interaction is predicted to increase accessibility of the Shine-Dalgarno sequence occluded in the long 5'-UTR and thus to promote Lmo0514 protein production inside the eukaryotic cell. Real-time quantitative PCR showed that Rli27 is upregulated in intracellular bacteria. In vivo experiments indicated a decrease in Lmo0514 protein levels in intracellular bacteria that lacked Rli27. Wild-type Lmo0514 levels were restored by expressing the wild-type Rli27 molecule but not a mutated version unable to interact with the lmo0514 long 5'-UTR. These findings emphasize how 5'-UTR length affects regulation by defined sRNA. In addition, they demonstrate how alterations in the relative abundance of two transcripts with distinct 5'-UTR confine the action of an sRNA for a specific target to bacteria that occupy the intracellular eukaryotic niche.

A novel antisense RNA from the Salmonella virulence plasmid pSLT expressed by non-growing bacteria inside eukaryotic cells.

Bacterial small RNAs (sRNAs) are regulatory molecules playing relevant roles in response to environmental changes, stressful conditions and pathogenesis. The intracellular bacterial pathogen Salmonella enterica serovar Typhimurium (S. Typhimurium) is known to regulate expression of some sRNAs during colonization of fibroblasts. Here, we characterize a previously unknown sRNA encoded in the S. Typhimurium pSLT virulence plasmid that is specifically up-regulated by non-growing dormant bacteria persisting inside fibroblasts. This sRNA was inferred in microarray expression analyses, which unraveled enhanced transcriptional activity in the PSLT047- PSLT046 (mig5) intergenic region. The sRNA transcript was further identified as a 597-nucleotide molecule, which we named IesR-1, for 'Intracellular-expressed-sRNA-1'. IesR-1 expression is low in bacteria growing in axenic cultures across a variety of experimental conditions but displays a marked increase (∼200-300 fold) following bacterial entry into fibroblasts. Remarkably, induction of IesR-1 expression is not prominent in bacteria proliferating within epithelial cells. IesR-1 deletion affects the control of bacterial growth in defined fibroblast cell lines and impairs virulence in a mouse infection model. Expression analyses performed in the PSLT047-iesR-1-PSLT046 (mig5) region support a cis-acting regulatory mechanism of IesR-1 as antisense RNA over the PSLT047 transcript involving interaction at their respective 3' ends and modulation of PSLT047 protein levels. This model is sustained by the scarce production of PSLT047 protein observed in non-growing intracellular bacteria and the high amount of PSLT047 protein produced by bacteria carrying a truncated IesR-1 version with separated 5' and 3' regions. Taken together, these data reveal that S. Typhimurium sRNAs encoded in the pSLT virulence plasmid respond to a state of persistence inside the host cell. As exemplified by IesR-1, some of these sRNAs may contribute to diminish the relative levels of proteins, such as PSLT047, which are probably dispensable for the intracellular lifestyle.

Genome expression analysis of nonproliferating intracellular Salmonella enterica serovar Typhimurium unravels an acid pH-dependent PhoP-PhoQ response essential for dormancy.

Genome-wide expression analyses have provided clues on how Salmonella proliferates inside cultured macrophages and epithelial cells. However, in vivo studies show that Salmonella does not replicate massively within host cells, leaving the underlying mechanisms of such growth control largely undefined. In vitro infection models based on fibroblasts or dendritic cells reveal limited proliferation of the pathogen, but it is presently unknown whether these phenomena reflect events occurring in vivo. Fibroblasts are distinctive, since they represent a nonphagocytic cell type in which S. enterica serovar Typhimurium actively attenuates intracellular growth. Here, we show in the mouse model that S. Typhimurium restrains intracellular growth within nonphagocytic cells positioned in the intestinal lamina propria. This response requires a functional PhoP-PhoQ system and is reproduced in primary fibroblasts isolated from the mouse intestine. The fibroblast infection model was exploited to generate transcriptome data, which revealed that ∼2% (98 genes) of the S. Typhimurium genome is differentially expressed in nongrowing intracellular bacteria. Changes include metabolic reprogramming to microaerophilic conditions, induction of virulence plasmid genes, upregulation of the pathogenicity islands SPI-1 and SPI-2, and shutdown of flagella production and chemotaxis. Comparison of relative protein levels of several PhoP-PhoQ-regulated functions (PagN, PagP, and VirK) in nongrowing intracellular bacteria and extracellular bacteria exposed to diverse PhoP-PhoQ-inducing signals denoted a regulation responding to acidic pH. These data demonstrate that S. Typhimurium restrains intracellular growth in vivo and support a model in which dormant intracellular bacteria could sense vacuolar acidification to stimulate the PhoP-PhoQ system for preventing intracellular overgrowth.

Dynamics of Salmonella small RNA expression in non-growing bacteria located inside eukaryotic cells.

Small non-coding regulatory RNAs (sRNAs) have been studied in many bacterial pathogens during infection. However, few studies have focused on how intracellular pathogens modulate sRNA expression inside eukaryotic cells. Here, we monitored expression of all known sRNAs of Salmonella enterica serovar Typhimurium (S. Typhimurium) in bacteria located inside fibroblasts, a host cell type in which this pathogen restrains growth. sRNA sequences known in S. Typhimurium and Escherichia coli were searched in the genome of S. Typhimurium virulent strain SL1344, the subject of this study. Expression of 84 distinct sRNAs was compared in extra- and intracellular bacteria. Non-proliferating intracellular bacteria upregulated six sRNAs, including IsrA, IsrG, IstR-2, RyhB-1, RyhB-2 and RseX while repressed the expression of the sRNAs DsrA, GlmZ, IsrH-1, IsrI, SraL, SroC, SsrS(6S) and RydC. Interestingly, IsrH-1 was previously reported as an sRNA induced by S. Typhimurium inside macrophages. Kinetic analyses unraveled changing expression patterns for some sRNAs along the infection. InvR and T44 expression dropped after an initial induction phase while IstR-2 was induced exclusively at late infection times (> 6 h). Studies focused on the Salmonella-specific sRNA RyhB-2 revealed that intracellular bacteria use this sRNA to regulate negatively YeaQ, a cis-encoded protein of unknown function. RyhB-2, together with RyhB-1, contributes to attenuate intracellular bacterial growth. To our knowledge, these data represent the first comprehensive study of S. Typhimurium sRNA expression in intracellular bacteria and provide the first insights into sRNAs that may direct pathogen adaptation to a non-proliferative state inside the host cell.

Human G3BP1 interacts with beta-F1-ATPase mRNA and inhibits its translation.

The post-transcriptional regulation of nuclear mRNAs that encode core components of mitochondria has relevant implications in cell physiology. The mRNA that encodes the catalytic subunit of the mitochondrial H(+)-ATP synthase subunit beta (ATP5B, beta-F1-ATPase) is localized in a large ribonucleoprotein (RNP) complex (beta-F1-RNP), which is subjected to stringent translational control during development and the cell cycle, and in carcinogenesis. Because downregulation of beta-F1-ATPase is a conserved feature of most prevalent human carcinomas, we have investigated the molecular composition of the human beta-F1-RNP. By means of an improved affinity-chromatography procedure and protein sequencing we have identified nine RNA-binding proteins (RNABPs) of the beta-F1-RNP. Immunoprecipitation assays of Ras-GAP SH3 binding protein 1 (G3BP1) and fluorescent in-situ hybridization of mRNA indicate a direct interaction of the endogenous G3BP1 with mRNA of beta-F1-ATPase (beta-F1 mRNA). RNA-bridged trimolecular fluorescence complementation (TriFC) assays confirm the interaction of G3BP1 with the 3'-UTR of beta-F1 mRNA in cytoplasmic RNA-granules. Confocal and high-resolution immunoelectron-microscopy experiments suggest that the beta-F1-RNP is sorted to the periphery of mitochondria. Molecular and functional studies indicate that the interaction of G3BP1 with beta-F1 mRNA inhibits its translation at the initiation level, supporting a role for G3BP1 in the glycolytic switch that occurs in cancer.

Up-regulation of the ATPase inhibitory factor 1 (IF1) of the mitochondrial H+-ATP synthase in human tumors mediates the metabolic shift of cancer cells to a Warburg phenotype.

The H(+)-ATP synthase is a reversible engine of mitochondria that synthesizes or hydrolyzes ATP upon changes in cell physiology. ATP synthase dysfunction is involved in the onset and progression of diverse human pathologies. During ischemia, the ATP hydrolytic activity of the enzyme is inhibited by the ATPase inhibitory factor 1 (IF1). The expression of IF1 in human tissues and its participation in the development of human pathology are unknown. Here, we have developed monoclonal antibodies against human IF1 and determined its expression in paired normal and tumor biopsies of human carcinomas. We show that the relative mitochondrial content of IF1 increases significantly in carcinomas, suggesting the participation of IF1 in oncogenesis. The expression of IF1 varies significantly in cancer cell lines. To investigate the functional activity of IF1 in cancer, we have manipulated its cellular content. Overexpression of IF1 or of its pH-insensitive H49K mutant in cells that express low levels of IF1 triggers the up-regulation of aerobic glycolysis and the inhibition of oxidative phosphorylation with concurrent mitochondrial hyperpolarization. Treatment of the cells with the H(+)-ATP synthase inhibitor oligomycin mimicked the effects of IF1 overexpression. Conversely, small interfering RNA-mediated silencing of IF1 in cells that express high levels of IF1 promotes the down-regulation of aerobic glycolysis and the increase in oxidative phosphorylation. Overall, these findings support that the mitochondrial content of IF1 controls the activity of oxidative phosphorylation mediating the shift of cancer cells to an enhanced aerobic glycolysis, thus supporting an oncogenic role for the de-regulated expression of IF1 in cancer.

Selective inhibition of beta-F1-ATPase mRNA translation in human tumours.

Down-regulation of beta-F1-ATPase (the catalytic subunit of the mitochondrial H+-ATP synthase) is a hallmark of many human tumours. The expression level of beta-F1-ATPase provides a marker of the prognosis of cancer patients, as well as of the tumour response to chemotherapy. However, the mechanisms that participate in down-regulating its expression in human tumours remain unknown. In the present study, we have investigated the expression of beta-F1-ATPase mRNA (termed beta-mRNA) in breast, colon and lung adenocarcinomas and squamous carcinomas of the lung. Despite the down-regulation of the protein, tumour beta-mRNA levels remained either unchanged (breast and lung adenocarcinomas) or significantly increased (colon and squamous lung carcinomas) when compared with paired normal tissues, suggesting a specific translation-masking event for beta-mRNA in human cancer. Consistently, we show using cell-free translation assays that a large fraction (approximately 70%) of protein extracts derived from breast and lung adenocarcinomas specifically repress the translation of beta-mRNA. We show that the 3'UTR (3' untranslated region) of human beta-mRNA is a relevant cis-acting element required for efficient translation of the transcript. However, an RNA chimaera bearing the 3'UTR of human beta-mRNA does not recapitulate the inhibitory effect of tumour extracts on beta-mRNA translation. Overall, the findings of the present study support the hypothesis that down-regulation of the bioenergetic activity of mitochondria in human tumours is exerted by translation silencing of beta-mRNA.

Cancer abolishes the tissue type-specific differences in the phenotype of energetic metabolism.

Nowadays, cellular bioenergetics has become a central issue of investigation in cancer biology. Recently, the metabolic activity of the cancer cell has been shown to correlate with a proteomic index that informs of the relative mitochondrial activity of the cell. Within this new field of investigation, we report herein the production and characterization of high-affinity monoclonal antibodies against proteins of the "bioenergetic signature" of the cell. The use of recombinant proteins and antibodies against the mitochondrial beta-F1-ATPase and Hsp60 proteins and the enzymes of the glycolytic pathway glyceraldehyde-3-phosphate dehydrogenase and pyruvate kinase M2 in quantitative assays provide, for the first time, the actual amount of these proteins in normal and tumor surgical specimens of breast, lung, and esophagus. The application of this methodology affords a straightforward proteomic signature that quantifies the variable energetic demand of human tissues. Furthermore, the results show an unanticipated finding: tumors from different tissues and/or histological types have the same proteomic signature of energetic metabolism. Therefore, the results indicate that cancer abolishes the tissue-specific differences in the bioenergetic phenotype of mitochondria. Overall, the results support that energetic metabolism represents an additional hallmark of the phenotype of the cancer cell and a promising target for the treatment of diverse neoplasias.

The tumor suppressor function of mitochondria: translation into the clinics.

Recently, the inevitable metabolic reprogramming experienced by cancer cells as a result of the onset of cellular proliferation has been added to the list of hallmarks of the cancer cell phenotype. Proliferation is bound to the synchronous fluctuation of cycles of an increased glycolysis concurrent with a restrained oxidative phosphorylation. Mitochondria are key players in the metabolic cycling experienced during proliferation because of their essential roles in the transduction of biological energy and in defining the life-death fate of the cell. These two activities are molecularly and functionally integrated and are both targets of commonly altered cancer genes. Moreover, energetic metabolism of the cancer cell also affords a target to develop new therapies because the activity of mitochondria has an unquestionable tumor suppressor function. In this review, we summarize most of these findings paying special attention to the opportunity that translation of energetic metabolism into the clinics could afford for the management of cancer patients. More specifically, we emphasize the role that mitochondrial beta-F1-ATPase has as a marker for the prognosis of different cancer patients as well as in predicting the tumor response to therapy.

Glucose avidity of carcinomas.

The cancer cell phenotype has been summarized in six hallmarks [D. Hanahan, R.A. Weinberg, The hallmarks of cancer, Cell 100 (1) (2000) 57-70]. Following the conceptual trait established in that review towards the comprehension of cancer, herein we summarize the basis of an underlying principle that is fulfilled by cancer cells and tumors: its avidity for glucose. Our purpose is to push forward that the metabolic reprogramming that operates in the cancer cell represents a seventh hallmark of the phenotype that offers a vast array of possibilities for the future treatment of the disease. We summarize the metabolic pathways that extract matter and energy from glucose, paying special attention to the concerted regulation of these pathways by the ATP mass-action ratio. The molecular and functional evidences that support the high glucose uptake and the "abnormal" aerobic glycolysis of the carcinomas are detailed discussing also the role that some oncogenes and tumor suppressors have in these pathways. We overview past and present evidences that sustain that mitochondria of the cancer cell are impaired, supporting the original Warburg's formulation that ascribed the high glucose uptake of cancer cells to a defective mitochondria. A simple proteomic approach designed to assess the metabolic phenotype of cancer, i.e., its bioenergetic signature, molecularly and functionally supports Warburg's hypothesis. Furthermore, we discuss the clinical utility that the bioenergetic signature might provide. Glycolysis is presented as the "selfish" pathway used for cellular proliferation, providing both the metabolic precursors and the energy required for biosynthetic purposes, in the context of a plethora of substrates. The glucose avidity of carcinomas is thus presented as the result of both the installment of glycolysis for cellular proliferation and of the impairment of mitochondrial activity in the cancer cell. At the end, the repression of mitochondrial activity affords the cancer cell with a cell-death resistant phenotype making them prone to malignant growth.

HuR and the bioenergetic signature of breast cancer: a low tumor expression of the RNA-binding protein predicts a higher risk of disease recurrence.

Downregulation of the catalytic subunit of the mitochondrial H(+)-ATP synthase (beta-F1-ATPase) is a hallmark of many types of cancer. The expression of beta-F1-ATPase is stringently controlled by posttranscriptional mechanisms. Herein, we pursue the identification of beta-F1-ATPase messenger RNA-binding proteins (beta-mRNABPs) that interact and could define the bioenergetic phenotype of the cancer cell in order to establish its relevance as markers of breast cancer progression. RNA immunoprecipitation and RNA affinity chromatography identify HuR as a beta-mRNABP that interacts with the 3'-untranslated region of the transcript. Subcellular fractionation and high-resolution immunoelectron microscopy revealed the cofractionation and presence of HuR in subcellular structures associated to liver mitochondria. Analysis of the expression level of HuR in a cohort of breast carcinomas shows its association with the degree of alteration of the bioenergetic phenotype of the tumor. Moreover, HuR expression is shown to be an independent marker of breast cancer prognosis. A low tumor expression of HuR predicts a higher risk of disease recurrence in early stage breast cancer patients as assessed by clinical and bioenergetic markers of prognosis, strongly supporting the incorporation of HuR as an additional marker for the follow-up of these patients. Mechanistically, overexpression experiments and short hairpin RNA-mediated silencing of HuR in human embryonic kidney and HeLa cells indicate that HuR is not regulating beta-F1-ATPase expression. Overall, the participation of additional RNA-binding proteins in controlling beta-F1-ATPase expression and therefore in defining the bioenergetic signature of the cancer cell is expected.

Loss of the mitochondrial bioenergetic capacity underlies the glucose avidity of carcinomas.

The down-regulation of the catalytic subunit of the mitochondrial H+-ATP synthase (beta-F1-ATPase) is a hallmark of most human carcinomas. This characteristic of the cancer cell provides a proteomic signature of cellular bioenergetics that can predict the prognosis of colon, lung, and breast cancer patients. Here we show that the in vivo tumor glucose uptake of lung carcinomas, as assessed by positron emission tomography in 110 patients using 2-deoxy-2-[18F]fluoro-d-glucose as probe, inversely correlates with the bioenergetic signature determined by immunohistochemical analysis in tumor surgical specimens. Further, we show that inhibition of the activity of oxidative phosphorylation by incubation of cancer cells with oligomycin triggers a rapid increase in their rates of aerobic glycolysis. Moreover, we show that the cellular expression level of the beta-F1-ATPase protein of mitochondrial oxidative phosphorylation inversely correlates (P < 0.001) with the rates of aerobic glycolysis in cancer cells. The results highlight the relevance of the alteration of the bioenergetic function of mitochondria for glucose capture and consumption by aerobic glycolysis in carcinomas.

A message emerging from development: the repression of mitochondrial beta-F1-ATPase expression in cancer.

Mitochondrial research has experienced a considerable boost during the last decade because organelle malfunctioning is in the genesis and/or progression of a vast array of human pathologies including cancer. The renaissance of mitochondria in the cancer field has been promoted by two main facts: (1) the molecular and functional integration of mitochondrial bioenergetics with the execution of cell death and (2) the implementation of (18)FDG-PET for imaging and staging of tumors in clinical practice. The latter, represents the bed-side translational development of the metabolic hallmark that describes the bioenergetic phenotype of most cancer cells as originally predicted at the beginning of previous century by Otto Warburg. In this minireview we will briefly summarize how the study of energy metabolism during liver development forced our encounter with Warburg's postulates and prompted us to study the mechanisms that regulate the biogenesis of mitochondria in the cancer cell.

Biogenesis and dynamics of mitochondria during the cell cycle: significance of 3'UTRs.

Nowadays, we are facing a renaissance of mitochondria in cancer biology. However, our knowledge of the basic cell biology and on the timing and mechanisms that control the biosynthesis of mitochondrial constituents during progression through the cell cycle of mammalian cells remain largely unknown. Herein, we document the in vivo changes on mitochondrial morphology and dynamics that accompany cellular mitosis, and illustrate the following key points of the biogenesis of mitochondria during progression of liver cells through the cycle: (i) the replication of nuclear and mitochondrial genomes is synchronized during cellular proliferation, (ii) the accretion of OXPHOS proteins is asynchronously regulated during proliferation being the synthesis of beta-F1-ATPase and Hsp60 carried out also at G2/M and, (iii) the biosynthesis of cardiolipin is achieved during the S phase, although full development of the mitochondrial membrane potential (DeltaPsim) is attained at G2/M. Furthermore, we demonstrate using reporter constructs that the mechanism regulating the accretion of beta-F1-ATPase during cellular proliferation is controlled at the level of mRNA translation by the 3'UTR of the transcript. The 3'UTR-driven synthesis of the protein at G2/M is essential for conferring to the daughter cells the original phenotype of the parental cell. Our findings suggest that alterations on this process may promote deregulated beta-F1-ATPase expression in human cancer.

Alteration of the bioenergetic phenotype of mitochondria is a hallmark of breast, gastric, lung and oesophageal cancer.

Recent findings indicate that the expression of the beta-catalytic subunit of the mitochondrial H+-ATP synthase (beta-F1-ATPase) is depressed in liver, kidney and colon carcinomas, providing further a bioenergetic signature of cancer that is associated with patient survival. In the present study, we performed an analysis of mitochondrial and glycolytic protein markers in breast, gastric and prostate adenocarcinomas, and in squamous oesophageal and lung carcinomas. The expression of mitochondrial and glycolytic markers varied significantly in these carcinomas, when compared with paired normal tissues, with the exception of prostate cancer. Overall, the relative expression of beta-F1-ATPase was significantly reduced in breast and gastric adenocarcinomas, as well as in squamous oesophageal and lung carcinomas, strongly suggesting that alteration of the bioenergetic function of mitochondria is a hallmark of these types of cancer.