The promyelocytic leukemia protein PML is organized into nuclear bodies which mediate suppression of oncogenic transformation and of growth. The biochemical functions of PML bodies are unknown, despite their involvement in several human disorders. We demonstrate that eukaryotic initiation factor 4E (eIF4E) directly binds the PML RING, a domain required for association with bodies and for suppression of transformation. Nuclear eIF4E functions in nucleocytoplasmic transport of a subset of transcripts including Cyclin D1. Present studies indicate that some PML requires the evolutionarily older eIF4E protein for association with nuclear bodies. Furthermore, PML RING modulates eIF4E activity by drastically reducing its af®nity for its substrate, 5¢ m 7 G cap of mRNA. We demonstrate that eIF4E requires cap binding for transport of Cyclin D1 mRNA and subsequent transformation activity. Additionally, PML reduces the af®nity of eIF4E for m 7 G mRNA cap, causing a reduction in Cyclin D1 protein levels and consequent transformation inhibition. PML is the ®rst factor shown to modulate nuclear eIF4E function. These ®ndings provide the ®rst biochemical framework for understanding the transformation suppression activity of PML. Keywords: eukaryotic initiation factor 4E/nuclear domain 10/promyelocytic leukemia/PML oncogenic domains/RING
Neurogenesis requires the coordination of neural progenitor proliferation and differentiation with cell-cycle regulation. However, the mechanisms coordinating these distinct cellular activities are poorly understood. Here we demonstrate for the first time that a Cut-like homeodomain transcription factor family member, Cux2 (Cutl2), regulates cell-cycle progression and development of neural progenitors. Cux2 loss-of-function mouse mutants exhibit smaller spinal cords with deficits in neural progenitor development as well as in neuroblast and interneuron differentiation. These defects correlate with reduced cell-cycle progression of neural progenitors coupled with diminished Neurod and p27Kip1 activity. Conversely, in Cux2 gain-of-function transgenic mice, the spinal cord is enlarged in association with enhanced neuroblast formation and neuronal differentiation, particularly with respect to interneurons. Furthermore, Cux2 overexpression induces high levels of Neurod and p27
Kip1. Mechanistically, we discovered through chromatin immunoprecipitation assays that Cux2 binds both the Neurod and p27Kip1 promoters in vivo, indicating that these interactions are direct. Our results therefore show that Cux2 functions at multiple levels during spinal cord neurogenesis. Cux2 initially influences cell-cycle progression in neural progenitors but subsequently makes additional inputs through Neurod and p27 Kip1 to regulate neuroblast formation, cell-cycle exit and cell-fate determination. Thus our work defines novel roles for Cux2 as a transcription factor that integrates cell-cycle progression with neural progenitor development during spinal cord neurogenesis.
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