No abstract

COP1 is a Ring-Finger E3 ubiquitin ligase that is involved in plant development, mammalian cell survival, growth, and metabolism. Here we report that COP1, whose expression is enhanced by insulin, regulates FoxO1 protein stability. We found that in Fao hepatoma cells, ectopic expression of COP1 decreased, whereas knockdown of COP1 expression increased the level of endogenous FoxO1 protein without impacting other factors such as C/EBP␣ and CREB (cAMP-response elementbinding protein). We further showed that COP1 binds FoxO1, enhances its ubiquitination, and promotes its degradation via the ubiquitin-proteasome pathway. To determine the biological significance of COP1-mediated FoxO1 protein degradation, we have examined the impact of COP1 on FoxO1-mediated gene expression and found that COP1 suppressed FoxO1 reporter gene as well as FoxO1 target genes such as glucose-6-phosphatase and phosphoenolpyruvate carboxykinase, two key targets for FoxO1 in the regulation of gluconeogenesis, with corresponding changes of hepatic glucose production in Fao cells. We suggest that by functioning as a FoxO1 E3 ligase, COP1 may play a role in the regulation of hepatic glucose metabolism.

Diffraction imaging of polystyrene spheres and B16F10 mouse melanoma cells embedded in gel has been investigated with a microscope objective. The diffraction images acquired with the objective from a sphere have been shown to be comparable to the Mie theory based projection images of the scattered light if the objective is translated to defocused positions towards the sphere. Using a confocal imaging based method to reconstruct and analyze the 3D structure, we demonstrated that genetic modifications in these cells can induce morphological changes and the modified cells can be used as an experimental model for study of the correlation between 3D morphology features and diffraction image data. Diffraction images of spheres (left column) and melanoma cells (right column) acquired with an objective.

Akt is activated on the plasma membrane and its substrates are distributed throughout various cellular compartments. To phosphorylate its substrates, Akt needs to be recruited to specific intracellular compartments. Thus, regulation of Akt cellular compartmentalization constitutes an important mechanism to specify Akt signaling. Here, we report the identification of ClipR-59 as an Akt interaction protein. We show that the interaction of ClipR-59 with Akt is mediated by the CAP-Gly domain of ClipR-59 and kinase domain of Akt and is regulated by Akt phosphorylation. We demonstrate that ClipR-59 regulates the Akt membrane association through its interaction with Akt and membrane localization and, by modulating Akt cellular compartmentalization, differentially modulates phosphorylation of Akt substrates in adipocytes. Finally, we provide evidence that one of the Akt substrates whose phosphorylation is upregulated by ClipR-59 is AS160, a negative regulator of adipocyte glucose transport. Accordingly, ectopic expression of ClipR-59 enhances, whereas knockdown of ClipR-59 suppresses, adipocyte glucose transport. We suggest that ClipR-59 functions as a scaffold protein that interacts with phospho-Akt and recruits active Akt on the membrane and may play an important role in adipocyte glucose transport.