Grasping Forkhead Box O functions by a bilateral approach

Metabolic adaptation to environmental alterations is fundamental for long term survival of the organism. Genetic and environmental manipulations proved that several signaling pathways are capable of regulating aging in response to environmental and physiological cues. The first example of a signaling module controlling longevity came from studies in Caenorhabditis elegans. Single mutations reducing insulin/IGF-1 signaling had a great potential to extend the lifespan of adult worms. Importantly, this effect turned out to be mediated by Forkhead Box O (FOXO) transcription factors, which become active upon inhibition of insulin/insulin growth factor 1 (IGF-1) signaling. The insulin/IGF-1/FOXO pathway is evolutionarily conserved and it appears now that it can also control the lifespan of higher organisms including humans. The capability of FOXO transcription factors to promote longevity is closely associated with their ability to respond to the elevated levels of reactive oxygen species (ROS) within the cell. This in turn results in transactivation of several antioxidants in a FOXO–dependent manner, which is associated with oxidative stress resistance. Previously, it has been shown that increased ROS levels induce several posttranslational modifications on FOXO proteins, including phosphorylation, acetylation and ubiquitination. Moreover, FOXOs specifically interact with other proteins in a ROS-dependent manner. Nevertheless, how FOXO activity is controlled under conditions of elevated ROS is still only partially understood. In this thesis, we have further investigated the molecular mechanisms underlying FOXO signaling in response to elevated cellular ROS and we focused our research on the FOXO family member FOXO4. We describe Nemo-like kinase (NLK) as a novel regulator of FOXO transcriptional activity. NLK binds to and phosphorylates FOXO1, FOXO3a, and FOXO4 on multiple residues. This co-occurs with inhibition of FOXO4 monoubiquitination. In the past, we have shown that oxidative stress-induced monoubiquitination of FOXO4 stimulates its nuclear retention. This correlates with increased transactivation of FOXO4, towards at least some of its target genes, and therefore might be important for the activation of antioxidant defensive programs. Conversely, NLK-dependent modulation of FOXO4 activity may also provide a means to downregulate this defensive program, when oxidative stress reaches a level beyond which repair is no longer feasible and cells need to undergo apoptosis. Furthermore, we identified Myb-binding protein 1a (MYBBP1a) as a novel interactor of FOXO4. Binding of MYBBP1a to FOXO4 is induced in response to elevated ROS levels and correlates with decreased acetylation of FOXO4. Since acetylation has been previously shown to inhibit FOXO activity, this consecutively promotes the activity of FOXO4 in a luciferase reporter assay. Interestingly, ectopic expression of MYBBP1a inhibits FOXO4-induced transactivation of the endogenous p27kip1 promoter, which might be the result of altered acetylation of histones. It has been indicated previously that association of FOXO with acetyl transferases can serve to acetylate histones and thereby open up the chromatin. Thus, in addition to decreasing the acetylation of FOXO4, MYBBP1a might hinder the acetylation of histones and thereby inhibit transcription. Previous in vivo studies revealed that FOXO proteins are redundant tumor suppressors, which might correlate with the promotion of longevity. However it becomes evident now that different FOXO proteins exhibit unique functions in the regulation of diverse physiological processes. Nevertheless, the understanding of the specific functions of FOXO4 is rather limited. Thus, to get a better insight into this issue we generated GFP-FOXO4 knock-in mice in order to examine the in vivo FOXO4 expression patterns. We were able to detect expression of FOXO4 only in progenitors of hematopoietic stem cells (HSCs) however the biological relevance of it needs to be determent

Activation of forkhead box O transcription factors by oncogenic BRAF promotes p21cip1-dependent senescence

Oncogene-induced senescence (OIS) is a potent tumor-suppressive mechanism that is thought to come at the cost of aging. The Forkhead box O (FOXO) transcription factors are regulators of life span and tumor suppression. However, whether and how FOXOs function in OIS have been unclear. Here, we show a role for FOXO4 in mediating senescence by the human BRAFV600E oncogene, which arises commonly in melanoma. BRAFV600E signaling through mitogen-activated protein kinase/extracellular signal-regulated kinase kinase resulted in increased reactive oxygen species levels and c-Jun NH 2 terminal kinase-mediated activation of FOXO4 via its phosphorylation on Thr223, Ser226, Thr447, and Thr451. BRAFV600E-induced FOXO4 phosphorylation resulted in p21cip1-mediated cell senescence independent of p16 ink4a or p27kip1. Importantly, melanocyte-specific activation of BRAFV600E in vivo resulted in the formation of skin nevi expressing Thr223/Ser226-phosphorylated FOXO4 and elevated p21cip1. Together, these findings support a model in which FOXOs mediate a trade-off between cancer and aging