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Most anticancer drugs entering clinical trials fail to achieve approval from the US FDA. Drug development is hampered by the lack of preclinical models with therapeutic predictive value. Herein, we report the development and validation of a tumorgraft model of renal cell carcinoma (RCC) and its application to the evaluation of an experimental drug. Tumor samples from 94 patients were implanted in the kidney of mice without additives or disaggregation. Tumors from 35 patients formed tumorgrafts, and 16 stable lines were established. Samples from metastatic sites engrafted at high frequency, and stable engraftment of primary tumors in mice correlated with decreased patient survival suggesting that tumor growth in mice may reveal the acquisition by the tumor of an ability to thrive at distant sites and metastasize. Tumorgrafts retained the histology, gene expression, DNA copy number alterations, and over 90% of the protein-coding gene mutations of the corresponding tumors. As determined by the induction of hypercalcemia in tumorgraft-bearing mice, tumorgrafts were able to act on the host causing paraneoplastic syndromes. In studies simulating drug exposures in patients, RCC tumorgraft growth was inhibited by sunitinib and sirolimus (into which temsirolimus is converted in humans), but not by erlotinib, which was used as a control. Dovitinib, a drug in clinical development, showed greater activity than sunitinib and sirolimus. The routine incorporation of models recapitulating the molecular genetics and drug sensitivities of human tumors into preclinical programs has the potential to improve oncology drug development.

Highlights d Pyruvate carboxylase (PC) maintains hepatic TCA cycle function and gluconeogenesis d PC replenishes TCA cycle intermediates required for urea cycle and redox capacity d Loss of hepatic PC prevents hyperglycemia in obesity but worsens liver inflammation d PC moderates oxidative stress during obesity by maintaining NADPH and glutathione

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Mitochondrial DNA is organized as a nucleoprotein complex called the nucleoid. Its major protein components have been identified in different organisms, but it is yet unknown whether nucleoids undergo any form of remodeling. Using an in organello ChIP-on-chip assay, we demonstrate that the DNA-bending protein Abf2 binds to most of the mitochondrial genome with a preference for GC-rich gene sequences. Thus, Abf2 is a bona fide mitochondrial DNA-packaging protein in vivo. Nucleoids form a more open structure under respiring growth conditions in which the ratio of Abf2 to mitochondrial DNA is decreased. Bifunctional nucleoid proteins Hsp60 and Ilv5 are recruited to nucleoids during glucose repression and amino-acid starvation, respectively. Thus, mitochondrial nucleoids in yeast are dynamic structures that are remodeled in response to metabolic cues. A mutant form of Hsp60 that causes mtDNA instability has altered submitochondrial localization, which suggests that nucleoid remodeling is essential for the maintenance of mitochondrial genome.

Mitochondrial function degenerates during aging and in aging-related neuromuscular degenerative diseases, which leads to the physiological decline of the cell 1 . Factors that can delay the degenerative process are actively sought after. Here, we show that reduced cytosolic protein synthesis is a robust cellular strategy that suppresses aging-related mitochondrial degeneration. We modelled the adultor later-onset degenerative disease, autosomal dominant Progressive External Ophthalmoplegia (adPEO), by introducing the A128P mutation into the yeast adenine nucleotide translocase, Aac2p. The aac2 A128P allele dominantly induces aging-dependent mitochondrial degeneration and phenotypically tractable degenerative cell death independent of its ADP/ATP exchange activity. Mitochondrial degeneration is suppressed by lifespan-extending nutritional interventions and by 8 longevity mutations, which are all known to reduce cytosolic protein synthesis. These longevity interventions also independently suppress aging-related mitochondrial degeneration in the pro-aging prohibitin mutants. The aac2 A128P mutant has reduced mitochondrial membrane potential (Δψ m ) and is synthetically lethal to low Δψ m conditions, including the loss of prohibitin. Mitochondrial degeneration is accelerated by defects in protein turnover on the inner membrane and is suppressed by cycloheximide, a specific inhibitor of cytosolic ribosomes. Reduced cytosolic protein synthesis suppresses membrane depolarization and defects in mitochondrial gene expression in aac2 A128P cells. Our finding thus provides a link between protein homeostasis (proteostasis), cellular bioenergetics and mitochondrial maintenance during aging.Autosomal dominant Progressive External Ophthalmoplegia (adPEO) is a neuromuscular degenerative disease clinically manifested by ptosis, progressive muscle weakness, sensory ataxia, peripheral neuropathy and parkinsonism 2 . Phenotypically, it is characterised by multiple deletions in mitochondrial DNA (mtDNA), while the activities of respiratory chain enzymes remain either normal or mildly affected 3 . One of the inherited forms of adPEO is caused by specific missense mutations in ANT1, encoding the isoform 1 of the adenine nucleotide translocase (Ant) 4 . Ant promotes ADP/ATP exchange across the mitochondrial inner membrane. Ant1 knock-out mice accumulate multiple mtDNA deletions in skeletal and cardiac muscles 5 . Low Ant activity depletes the matrix ADP level. This sequentially causes ATP synthase stagnation, membrane hyperpolarization, increased free radical production and mtDNA damages. Interestingly, mutations in the yeast Aac2p, that mimic the pathogenic 3Correspondence should be addressed to X.J.C. (chenx@upstate.edu). Note: Supplementary Information is available on the Nature Cell Biology website.Author Information: The authors declare no competing financial interests. X.W., X.Z., B.K. and X.J.C. performed experiments; X.J.C. designed experiments, analyzed data and wrote the manuscript. All authors discussed the results a...

Mammalian target of rapamycin complex 1 (mTORC1) is implicated in cell growth control and is extensively regulated. We previously reported that in response to hypoxia, mTORC1 is inhibited by the protein regulated in development and DNA damage response 1 (REDD1). REDD1 is upregulated by HIF-1, and forced REDD1 expression is sufficient to inhibit mTORC1. REDD1-induced mTORC1 inhibition is dependent on a protein complex formed by the tuberous sclerosis complex (TSC)1 and 2 (TSC2) proteins. In clear-cell renal cell carcinoma (ccRCC), the von Hippel-Lindau (VHL) gene is frequently inactivated leading to constitutive activation of HIF-2 and/or HIF-1, which may be expected to upregulate REDD1 and inhibit mTORC1. However, mTORC1 is frequently activated in ccRCC and mTORC1 inhibitors are effective against this tumor type; a paradox herein examined. REDD1 was upregulated in VHL-deficient ccRCC by in silico microarray analyses, as well as by quantitative real-time PCR, Western blot, and immunohistochemistry. Vhl disruption in a mouse model was sufficient to induce Redd1. Using ccRCC-derived cell lines, we show that REDD1 upregulation in tumors is VHL-dependent, and that both HIF-1 and HIF-2 are, in a cell-type dependent manner, recruited to, and essential for, REDD1 induction. Interestingly, whereas mTORC1 is responsive to REDD1 in some tumors, strategies have evolved in others, such as mutations disrupting TSC1, to subvert mTORC1 inhibition by REDD1. Sequencing analyses of 77 ccRCCs for mutations in TSC1, TSC2 and REDD1, using PTEN as a reference, implicate the TSC1 gene, and possibly REDD1, as tumor suppressors in sporadic ccRCC. Understanding how ccRCCs become refractory to REDD1-induced mTORC1 inhibition should shed light into the development of ccRCC and may aid in patient selection for molecular targeted therapies.