Dissecting the signaling events that impact classical nuclear import and target nuclear transport factors.
BACKGROUND
Signaling through MEK-->ERK1/2 and PI3 kinases is implicated in many aspects of cell physiology, including the survival of oxidant exposure. Oxidants play a role in numerous physiological and pathophysiological processes, many of which rely on transport in and out of the nucleus. However, how oxidative stress impacts nuclear trafficking is not well defined.


RESULTS
To better understand the effect of stress on nucleocytoplasmic trafficking, we exposed cells to the oxidant diethyl maleate. This treatment activated MEK-->ERK1/2 as well as PI3 kinase-->Akt cascades and triggered the inhibition of classical nuclear import. To define the molecular mechanisms that regulate nuclear transport, we examined whether MEK and PI3 kinase signaling affected the localization of key transport factors. Using recently developed tools for image acquisition and analysis, the subcellular distributions of importin-alpha, CAS, and nucleoporins Nup153 and Nup88 were quantified in different cellular compartments. These studies identified specific profiles for the localization of transport factors in the nucleus and cytoplasm, and at the nuclear envelope. Our results demonstrate that MEK and PI3 kinase signaling as well as oxidative stress control nuclear trafficking and the localization of transport components. Furthermore, stress not only induced changes in transport factor distribution, but also upregulated post-translational modification of transport factors. Our results are consistent with the idea that the phosphorylation of importin-alpha, CAS, Nup153, and Nup88, and the O-GlcNAc modification of Nup153 increase when cells are exposed to oxidant.


CONCLUSIONS
Our studies defined the complex regulation of classical nuclear import and identified key transport factors that are targeted by stress, MEK, and PI3 kinase signaling.
Kodiha
Mohamed
M
Department of Physiology, McGill University, Montreal, Canada.
Tran
Dan
D
Morogan
Andreea
A
Qian
Cynthia
C
Stochaj
Ursula
U
D021581:Active Transport, Cell Nucleus / Q000187:drug effects; D002451:Cell Compartmentation / Q000187:drug effects; D002467:Cell Nucleus / Q000378:metabolism*; D029563:Cellular Apoptosis Susceptibility Protein / Q000378:metabolism; D017930:Genes, Reporter; D006367:HeLa Cells; D006801:Humans; D020935:MAP Kinase Signaling System / Q000187:drug effects; D020929:Mitogen-Activated Protein Kinase Kinases / Q000037:antagonists & inhibitors; D009685:Nuclear Envelope / Q000378:metabolism; D019913:Nuclear Localization Signals / Q000378:metabolism; D028861:Nuclear Pore Complex Proteins / Q000378:metabolism; D009687:Nuclear Proteins / Q000378:metabolism*; D018384:Oxidative Stress / Q000187:drug effects; D000081082:Phosphoinositide-3 Kinase Inhibitors; D047428:Protein Kinase Inhibitors / Q000494:pharmacology; D011499:Protein Processing, Post-Translational / Q000187:drug effects; D015398:Signal Transduction / Q000187:drug effects; D012995:Solubility / Q000187:drug effects; D013312:Stress, Physiological / Q000187:drug effects; D015854:Up-Regulation / Q000187:drug effects; D028901:alpha Karyopherins / Q000378:metabolism
D029563:Cellular Apoptosis Susceptibility Protein; D019913:Nuclear Localization Signals; D028861:Nuclear Pore Complex Proteins; D009687:Nuclear Proteins; D000081082:Phosphoinositide-3 Kinase Inhibitors; D047428:Protein Kinase Inhibitors; D028901:alpha Karyopherins; D020929:Mitogen-Activated Protein Kinase Kinases