Exportin 7 (RanBP16) Plays An Essential Role in Terminal Erythroid Chromatin Condensation and Enucleation

Abstract 178

In contrast to compacting chromatin into highly condensed mitotic chromosomes, the quite distinct process of global chromatin condensation culminating in enucleation that occurs during terminal erythroid development is still poorly understood. By examining the protein composition of the erythroid nucleus from early erythroblast to ultimate extrusion, I observed that extruded nuclei are largely depleted of all nuclear proteins. Given my previous observations that the highly-regulated but reportedly nonspecific nuclear export protein, Xpo7 or RanBP16, is highly induced during erythropoiesis and highly erythroid-specific, I hypothesized that its role may be to remove almost all nuclear proteins in order to allow the terminal erythroid chromatin to condense. Knockdown of Xpo7 using shRNA in primary fetal liver erythroid progenitors resulted in severe inhibition of chromatin condensation and enucleation but had little effect on hemoglobin accumulation or erythroid cell surface marker expression. As expected based on my hypothesis, proteomic analysis of nuclei from Xpo7-knockdown cells revealed largely all nuclear proteins, some of which may be responsible for the process of histone redistribution during chromatin condensation. Xpo7 is also highly regulated: besides its promoter being bound directly by the erythroid master regulators GATA1 and Klf1 (unpublished data), it is also the target of a miRNA whose level decreases during erythropoiesis, miR-181, and whose overexpression has been shown to result in the inhibition of terminal enucleation. Because chromatin condensation occurs in lower vertebrates without subsequent enucleation, I have also explored the localization and function of Xpo7 in zebrafish using in situ hybridization and morpholinos, respectively, and found that this export function is specific to mammalian chromatin condensation, providing evidence that condensation and enucleation are inextricably linked processes in mammals.