The small GTPases Rac1 and Rac2 regulate mDia2 to promote asymmetric cell division and enucleation in developing erythrocytes.
A critical step in the formation of mature mammalian erythrocytes from progenitor cells is an asymmetric cell division in which the nucleus is compacted, extruded and enveloped by the plasma membrane. The enucleated daughter cell, known as a reticulocyte, eventually matures into an erythrocyte. While other facets of erythropoeisis have been extensively characterized, the signaling pathways that regulate enucleation remain poorly understood. In Nature Cell Biology, Lodish and colleagues now show that the Rac1 and Rac2 GTPases signal through mDia2 to promote enucleation in mouse erythrocytes.
Erythropoeisis can be observed in vitro by harvesting erythroid progenitor cells from E13.5 embryonic mouse livers and culturing them with erythropoietin and fibronectin. Previous studies have indicated that actin filaments form a cortical actin ring (CAR) between the condensed nucleus and the incipient reticulocyte. The CAR then contracts and pinches the nucleus off to form a reticulocyte. Rho family GTPases are well-known regulators of the actin cytoskeleton and govern CAR formation during cytokinesis. To evaluate a role for Rho proteins in enucleation, erythroid progenitor cells were infected with dominant-negative Rac, Rho and Cdc42. Dominant-negative RhoA and Cdc42 had very little effect on enucleation, whereas pharmacologic inhibition of Rac, or expression of dominant-negative Rac1 or Rac2, blocked both formation of the CAR and enucleation. Rac-deficient cells displayed no other defects in cell proliferation or differentiation.
The RhoA effector mDia nucleates the CAR during cytokinesis. Interestingly, mDia2 and Rac GTPases were found to localize to the CAR during enucleation. Short hairpin RNA-mediated downregulation of mDia2 blocked enucleation and produced a phenotype similar to Rac-deficient cells. Furthermore, Rac1 and Rac2 interacted with mDia2 in a GTP-dependent manner in vitro, suggesting that mDia2 is involved in Rac-mediated enucleation. Indeed, expression of constitutively active mDia2 rescued the enucleation defect observed upon expression of dominant-negative Rac1 or Rac2. It remains unclear if mDia2 is a direct Rac effector, as a robust direct interaction could not be verified in vivo; nonetheless, Rac–mDia2 signaling is required for formation of the CAR and enucleation.
These data are consistent with the idea that small GTPases are required for the establishment of polarity and asymmetric cell division in vertebrates, and provide the first insights into how enucleation of erythroid progenitor cells is regulated at the molecular level. Future studies will be important to understand the signals that regulate Rac and promote enucleation in immature mammalian erythroblasts.
Emily J. Chenette Signaling Gateway
Reference
Ji, P., Jayapal, S. R. & Lodish, H. F. Enucleation of cultured mouse fetal erythroblasts requires Rac GTPases and mDia2. Nature Cell Biology10, 314-321 (2008) | Article | PubMed |
