Lamin B1 mediates dynamic changes in heterochromatin and regulates erythropoiesis

Abstract Chromatin condensation is a crucial step in the maturation process of red blood cells. we explored the distribution changes of chromatin modifications during erythroid differentiation, the dynamic regulatory factors of Lamin B1, and their relationship with changes in chromatin structure. Using an in vitro erythroid differentiation system, we isolated six distinct differentiation stages, CFU-E, proerythroblast (ProE), basophilic erythroblast (EB), late basophilic erythroblast (LB), polychromatophilic erythroblast (Poly), and orthochromatic erythroblast (Ortho)via fluorescence-activated cell sorting (FACS). We employed BL-Hi-C, ChIP-seq, immunofluorescence, and chromosome painting to investigate chromatin conformation changes during erythroid development. Hi-C analysis revealed that chromatin compaction initiates at the LB stage. Moreover, increased long-range chromatin interactions in late erythropoiesis are predominantly driven by B compartment. ChIP assays targeting histone modifications revealed that the B compartment is enriched with H3K9me3, a canonical heterochromatin mark, whose levels progressively increase during erythroid differentiation. Western blotting and immunofluorescence analyses of H3K9me3 showed no significant global increase in total protein levels during differentiation. However, we observed a marked spatial redistribution of H3K9me3, which closely correlates with DNA condensation and exhibits extensive co-localization with the nuclear membrane at later stages. From CFU-E to Ortho stages, H3K9me3 enrichment at genic regions remains largely unchanged. In contrast, a subset of intergenic regions shows significant H3K9me3 accumulation starting at the late basophilic stage, overlapping with B compartments and lamina-associated domains (LADs). At the late basophilic stage, lamina-associated domains (LADs) undergo structural disruption, accompanied by significant reductions in both genomic coverage and domain length. Immunofluorescence further demonstrated dissociation of Lamin B1 from the nuclear membrane, enabling the release of peripheral chromatin into the nucleoplasm. This redistribution enhances chromatin mobility and facilitates large-scale chromatin compaction. Our findings indicate that Lamin B1 plays a central role in regulating heterochromatin dynamics during erythropoiesis. During the early stages of erythroid differentiation, Lamin B1 ensures a normal differentiation process. In the late basophilic stage, the absence of Lamin B1 leads to the disruption of Lamina-Associated Domains (LADs), which in turn promotes changes in higher-order chromatin structure mediated by heterochromatin. That is, at different stages of erythroid differentiation, the nuclear Lamina protein Lamin B1 exerts precise regulatory control and is an important regulatory factor for higher-order chromatin structure.