George Thomas

Humans have evolved elaborate mechanisms to activate p53 in response to insults that lead to cancer, including the binding and inhibition of Hdm2 by the 60S ribosomal proteins (RPs) RPL5 and RPL11. This same mechanism appears to be activated upon impaired ribosome biogenesis, a risk factor for cancer initiation. As loss of RPL5/RPL11 abrogates ribosome biogenesis and protein synthesis to the same extent as loss of other essential 60S RPs, we reasoned the loss of RPL5 and RPL11 would induce a p53-independent cell cycle checkpoint. Unexpectedly, we found that their depletion in primary human lung fibroblasts failed to induce cell cycle arrest but strongly suppressed cell cycle progression. We show that the effects on cell cycle progression stemmed from reduced ribosome content and translational capacity, which suppressed the accumulation of cyclins at the translational level. Thus, unlike other tumor suppressors, RPL5/RPL11 play an essential role in normal cell proliferation, a function cells have evolved to rely on in lieu of a cell cycle checkpoint.

Hematopoietic stem/progenitor cells (HSPCs) function to give rise to mature blood cells. Effective DNA damage response (DDR) and maintenance of genomic stability are crucial for normal functioning of HSPCs. Mammalian target of rapamycin (mTOR) integrates signals from nutrients and growth factors to control protein synthesis, cell growth, survival and metabolism, and has been shown to regulate DDR in yeast and human cancer cells through the p53/p21 signaling cascade. Here, we show that gene targeting of mTOR in HSPCs causes a defective DDR due to a variety of DNA damage agents, mimicking that caused by deficient FANCD2, a key component of the Fanconi anemia (FA) DDR machinery. Mechanistically, mTOR^−/− HSPCs express drastically reduced FANCD2. Consistent with these genetic findings, inactivation of mTOR in human lymphoblast cells by pp242 or Torin 1, mTOR kinase inhibitors, suppresses FANCD2 expression and causes a defective DDR that can be rescued by reconstitution of exogenous FANCD2. Further mechanistic studies show that mTOR deficiency or inactivation increases phosphorylation and nuclear translocation of nuclear factor (NF)-κB, which results in an enhanced NF-κB binding to FANCD2 promoter to suppress FANCD2 expression. Thus, mTOR regulates DDR and genomic stability in hematopoietic cells through a noncanonical pathway involving NF-κB-mediated FANCD2 expression.

Mechanistic target of rapamycin (Mtor) is required for embryonic inner cell mass proliferation during early development. However, Mtor expression levels are very low in the mouse heart during embryogenesis. To determine if Mtor plays a role during mouse cardiac development, cardiomyocyte specific Mtor deletion was achieved using α myosin heavy chain (α-MHC) driven Cre recombinase. Initial mosaic expression of Cre between embryonic day (E) 10.5 and E11.5 eliminated a subset of cardiomyocytes with high Cre activity by apoptosis and reduced overall cardiac proliferative capacity. The remaining cardiomyocytes proliferated and expanded normally. However loss of 50% of cardiomyocytes defined a threshold that impairs the ability of the embryonic heart to sustain the embryo’s circulatory requirements. As a result 92% of embryos with cardiomyocyte Mtor deficiency died by the end of gestation. Thus Mtor is required for survival and proliferation of cardiomyocytes in the developing heart.

Combination therapy causes gene reprogramming, autophagy, and tumor regression in a mouse model approximating human HCC.

Impairment of ribosome biogenesis leads to p53 induction and cell cycle arrest, a checkpoint involved in human disease. Induction of p53 is attributed to the binding and inhibition of human double minute 2 (Hdm2) by a subset of ribosomal proteins (RPs): RPS7, RPL5, RPL11, and RPL23. However, we found that only RPL11 or RPL5, in a mutually dependent manner, elicit this response. We show that depletion of RPS7 or RPL23, like depletion of other RPs, except for RPL11 and RPL5, induces a p53 response and that the effects of RPS7 and RPL23 on p53 induction reported earlier may be ascribed to inhibition of global translation. Moreover, we made the surprising observation that codepletion of two essential RPs, one from each subunit, but not the same subunit, leads to suprainduction of p53. This led to the discovery that the previously proposed RPL11-dependent mechanism of p53 induction, thought to be caused by abrogation of 40S biogenesis and continued 60S biogenesis, is still operating, despite abrogation of 60S biogenesis. This response leads to both a G1 block and a novel G2/M block not observed when disrupting either subunit alone. Thus, induction of p53 is mediated by distinct mechanisms, with the data pointing to an essential role for ribosomal subunits beyond translation.

Activation of PLD1 by amino acids enables mTORC1 to respond to nutrient abundance.

Targeted deletion of S6 kinase (S6K) 1 in mice leads to higher energy expenditure and improved glucose metabolism. However, the molecular mechanisms controlling these effects remain to be fully elucidated. Here, we analyze the potential role of dietary lipids in regulating the mTORC1/S6K system. Analysis of S6K phosphorylation in vivo and in vitro showed that dietary lipids activate S6K, and this effect is not dependent upon amino acids. Comparison of male mice lacking S6K1 and 2 (S6K-dko) with wt controls showed that S6K-dko mice are protected against obesity and glucose intolerance induced by a high-fat diet. S6K-dko mice fed a high-fat diet had increased energy expenditure, improved glucose tolerance, lower fat mass gain, and changes in markers of lipid metabolism. Importantly, however, these metabolic phenotypes were dependent upon dietary lipids, with no such effects observed in S6K-dko mice fed a fat-free diet. These changes appear to be mediated via modulation of cellular metabolism in skeletal muscle, as shown by the expression of genes involved in energy metabolism. Taken together, our results suggest that the metabolic functions of S6K in vivo play a key role as a molecular interface connecting dietary lipids to the endogenous control of energy metabolism.

Comment on: Morgado-Palacin L, et al. Cell Cycle 2012; 11:503–10