Dynamic subcellular partitioning of the nucleolar transcription factor TIF-IA under ribotoxic stress

作者：Szymanski Jedrzej; Mayer Christine; Hoffmann Rohrer Urs; Kalla Claudia; Grummt Ingrid; Weiss Matthias^*

来源：Biochimica et Biophysica Acta-Molecular Cell Research, 2009, 1793(7): 1191-1198.

DOI：10.1016/j.bbamcr.2009.05.004

• Nucleolus
• Ribotoxic stress
• TIF-IA
• Fluorescence recovery after photobleaching
• Fluorescence correlation spectroscopy

TIF-IA is a basal transcription factor of RNA polymerase I (Pol I) that is a major target of the JNK2 signaling pathway in response to ribotoxic stress. Using advanced fluorescence microscopy and kinetic modeling we elucidated the subcellular localization of TIF-IA and its exchange dynamics between the nucleolus, nucleoplasm and cytoplasm upon ribotoxic stress. In steady state, the majority of (GFP-tagged) TIF-IA was in the cytoplasm and the nucleus, a minor portion (7%) localizing to the nucleoli. We observed a rapid shuttling of GFP-TIF-IA between the different cellular compartments with a mean residence time of similar to 130 s in the nucleus and only similar to 30 s in the nucleoli. The import rate from the cytoplasm to the nucleus was similar to 3-fold larger than the export rate, suggesting an importin/exportin-mediated transport rather than a passive diffusion. Upon ribotoxic stress, GFP-TlF-IA was released from the nucleoli with a half-time of similar to 24 min. Oxidative stress and inhibition of protein synthesis led to a relocation of GFP-TIF-IA with slower kinetics while osmotic stress had no effect. The observed relocation was much slower than the nucleo-cytoplasmic and nucleus-nucleolus exchange rates of GFP-TIF-IA, indicating a time-limiting step upstream of the JNK2 pathway. In support of this, time-course experiments on the activity of JNK2 revealed the activation of the JNK kinase as the rate-limiting step.

• 出版日期2009-7