Unicellular organisms such as yeasts require a single cyclin-dependent kinase, Cdk1, to drive cell division. In contrast, mammalian cells are thought to require the sequential activation of at least four different cyclin-dependent kinases, Cdk2, Cdk3, Cdk4 and Cdk6, to drive cells through interphase, as well as Cdk1 to proceed through mitosis. This model has been challenged by recent genetic evidence that mice survive in the absence of individual interphase Cdks. Moreover, most mouse cell types proliferate in the absence of two or even three interphase Cdks. Similar results have been obtained on ablation of some of the activating subunits of Cdks, such as the D-type and E-type cyclins. Here we show that mouse embryos lacking all interphase Cdks (Cdk2, Cdk3, Cdk4 and Cdk6) undergo organogenesis and develop to midgestation. In these embryos, Cdk1 binds to all cyclins, resulting in the phosphorylation of the retinoblastoma protein pRb and the expression of genes that are regulated by E2F transcription factors. Mouse embryonic fibroblasts derived from these embryos proliferate in vitro, albeit with an extended cell cycle due to inefficient inactivation of Rb proteins. However, they become immortal on continuous passage. We also report that embryos fail to develop to the morula and blastocyst stages in the absence of Cdk1. These results indicate that Cdk1 is the only essential cell cycle Cdk. Moreover, they show that in the absence of interphase Cdks, Cdk1 can execute all the events that are required to drive cell division.

Amyotrophic lateral sclerosis (ALS) is a rapidly progressing neurodegenerative disease, characterized by motor neuron (MN) death, for which there are no truly effective treatments. Here, we describe a new small molecule survival screen carried out using MNs from both wildtype and mutant SOD1 mouse embryonic stem cells. Among the hits we found, kenpaullone had a particularly impressive ability to prolong the healthy survival of both types of MNs that can be attributed to its dual inhibition of GSK3 and HGK kinases. Furthermore, kenpaullone also strongly improved the survival of human MNs derived from ALS patient induced pluripotent stem cells and was more active than either of two compounds, olesoxime and dexpramipexole, that recently failed in ALS clinical trials. Our studies demonstrate the value of a stem cell approach to drug discovery and point to a new paradigm for identification and preclinical testing of future ALS therapeutics.

Mammalian cell division is thought to be driven by sequential activation of several Cyclin-dependent kinases (Cdk), mainly Cdk4, Cdk6, Cdk2 and Cdk1. Since mice lacking Cdk4, Cdk6 or Cdk2 are viable, it has been proposed that they play compensatory roles. We report here that mice lacking Cdk4 and Cdk2 complete embryonic development to die shortly thereafter presumably due to heart failure. However, conditional ablation of Cdk2 in adult mice lacking Cdk4 does not result in obvious abnormalities. Moreover, these double mutant mice recover normally after partial hepatectomy. In culture, Cdk4(-/-);Cdk2(-/-) embryonic fibroblasts become immortal, display robust pRb phosphorylation and have normal S phase kinetics. These observations indicate that Cdk4 and Cdk2 are dispensable for the mammalian cell cycle and for adult homeostasis.

The motor neuron disease Spinal Muscular Atrophy (SMA) results from mutations that lead to low levels of the ubiquitously expressed protein Survival of Motor Neuron (SMN). Ever-increasing data suggest that therapeutics that elevate SMN may be effective in treating SMA. We executed an image-based screen of annotated chemical libraries and discovered multiple classes of compounds that were able to increase cellular SMN. Among the most important was the RTK/PI3K/AKT/GSK-3 signaling cascade. Chemical inhibitors of GSK-3, as well as shRNAs directed against this target, elevate SMN levels primarily by stabilizing the protein. Of particular significance is that GSK-3 chemical inhibitors were also effective in motor neurons, not only in elevating SMN levels, but also in blocking the death that was produced when SMN was acutely reduced by a SMN-specific shRNA. Thus, we have established a screen capable of detecting drug-like compounds that correct the main phenotypic change that underlies SMA.