The Nup84 nuclear pore subcomplex mediates segregation of the opposing Rap1 roles in transcriptional repression and activation.
Balaraj Menon (1), Nayan Sarma (1), Satish Pasula (1), Stephen Deminoff (2), Kristine Willis (3), Kellie Barbara (1), Brenda Andrews (3), George Santangelo (3)
(1) Department of Biological Sciences, University of Southern Mississippi, 118 College Drive, Hattiesburg, MS 39406;
(2) Department of Molecular Genetics, The Ohio State University, 484 W. 12th Ave., Columbus, Ohio 43210;
(3) Medical Genetics, University of Toronto, 1 Kings College Cir., Toronto, ON, M5S 1A8, Canada
An important goal in modern biology is to understand how activator/repressor proteins (e.g. p53, MyoD and HoxA9 in mammalian systems) simultaneously participate in opposite regulatory roles. A prominent example in yeast is the multifunctional regulator Rap1, which recognizes identical motifs in mediating either repression (at silent mating type loci and at telomeres) or activation (at promoters of glycolytic genes and ribosomal protein genes). We show here that the Rap1 activation assemblage, like its silencing complex, is anchored at the nuclear periphery through attachment to nucleoporins and associated factors. We used multiple independent approaches to establish this physical and functional link between Rap1 activation and nuclear pores. We did a triple fluor experiment (YFP+CFP+RFP) to visualize Rap1 activation complexes at the nuclear periphery. We used synthetic genetic array (SGA) analysis to identify a robust genetic network that connects Rap1 activation with the Nup84 subcomplex and nuclear envelope components. We also co-immunoprecipitated these same nuclear pore factors with Rap1 co-activators. Finally, we found that a subset of nucleoporins activate transcription when fused to a DNA binding domain, a result that recapitulates oncogenic nucleoporin-HoxA9 gene fusions found in acute leukemias in humans. Nup84 activation is entirely dependent on Rap1 co-factors. The implications of this discovery for understanding gene regulation in eukaryotes will be discussed.
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