Mammalian red blood cells lack nuclei. The molecular mechanisms underlying erythroblast nuclear condensation and enucleation, however, remain poorly understood. Here we show that Wdr26, a gene upregulated during terminal erythropoiesis, plays an essential role in regulating nuclear condensation in differentiating erythroblasts. Loss of Wdr26 induces anemia in zebrafish and enucleation defects in mouse erythroblasts because of impaired erythroblast nuclear condensation. As part of the glucose-induced degradation-deficient ubiquitin ligase complex, Wdr26 regulates the ubiquitination and degradation of nuclear proteins, including lamin B. Failure of lamin B degradation blocks nuclear opening formation leading to impaired clearance of nuclear proteins and delayed nuclear condensation. Collectively, our study reveals an unprecedented role of an E3 ubiquitin ligase in regulating nuclear condensation and enucleation during terminal erythropoiesis. Our results provide mechanistic insights into nuclear protein homeostasis and vertebrate red blood cell development.

Developing erythroblasts acquire massive amounts of iron through the transferrin (Tf) cycle, which involves endocytosis, sorting, and recycling of the Tf-Tf receptor (Tfrc) complex. Previous studies on the hemoglobin-deficit (hbd) mouse have shown that the exocyst complex is indispensable for the Tfrc recycling, however, the precise mechanism underlying the efficient exocytosis and recycling of Tfrc in erythroblasts remains unclear. Here we identify the guanine nucleotide exchange factor Grab as a critical regulator of the Tf cycle and iron metabolism during erythropoiesis. Grab is highly expressed in differentiating erythroblasts. Loss of Grab diminishes the Tfrc recycling and iron uptake, leading to hemoglobinization defects in mouse primary erythroblasts, mammalian erythroleukemia cells, and zebrafish embryos. These defects can be alleviated by supplementing iron together with hinokitiol, a small-molecule natural compound that can mediate iron transport independent of the Tf cycle. Mechanistically, Grab regulates the exocytosis of Tfrc-associated vesicles by activating the GTPase Rab8, which subsequently promotes the recruitment of the exocyst complex and vesicle exocytosis. Our results reveal a critical role for Grab in regulating the Tf cycle and provide new insights into iron homeostasis and erythropoiesis.

Vesicle trafficking is a fundamental cellular process that controls the transport of various proteins and cargos between cellular compartments in eukaryotes. Using a combination of genome‐wide CRISPR screening in mammalian cells and RNAi screening in Caenorhabditis elegans, we identify chaperonin containing TCP‐1 subunit 4 (CCT4) as a critical regulator of protein secretion and vesicle trafficking. In C. elegans, deficiency of cct‐4 as well as other CCT subunits impairs the trafficking of endocytic markers in intestinal cells, and this defect resembles that of dyn‐1 RNAi worms. Consistent with these findings, the silencing of CCT4 in human cells leads to defective endosomal trafficking, and this defect can be rescued by the dynamin activator Ryngo 1–23. These results suggest that the cytosolic chaperonin CCT may regulate vesicle trafficking by promoting the folding of dynamin in addition to its known substrate tubulin. Our findings establish an essential role for the CCT chaperonin in regulating vesicle trafficking, and provide new insights into the regulation of vesicle trafficking and the cellular function of the cytosolic chaperonin.