Hematopoietic stem cells, essential for the production of blood cells, decline in function with aging. Among these changes with aging are increased frequency of mutations in epigenetic modifiers. We analyzed the epigenetic landscape of Lineage-CD34+CD38- human HSC-enriched fraction (HSCe) from young (18-30 years), middle-aged (45-55 years), and aged (65-75 years) individuals by comparing changes in gene expression by RNA sequencing and histone marks (H3K4me3, H3K4me1, H3K27ac, H3K27me3) by ChIP sequencing. Integrative analysis of the histone profiles by k-means clustering identified 12 clusters of genes defined by their specific pattern of age-associated epigenetic changes, which were accompanied by changes in gene expression affecting 1,133 genes between young and aged HSCe. The LMNA gene, which encodes the nuclear lamina protein Lamin A/C, was one of the most downregulated genes with aging (7.9-fold, p=1.9x10-13). This downregulation was mediated by loss of active histone marks H3K4me3 at the LMNA gene promoter, as well as loss of H3K27ac at two putative enhancer regions. ShRNA-mediated knockdown of LMNA in young, CD34+ hematopoietic stem and progenitor cells (HSPCs) resulted in an increase in myeloid colony formation (n=7 biological replicates with an average of 33% in total colony increase), consistent with the aging phenotype. Moreover, knockdown of LMNA in CD34+ cells impaired myeloid differentiation in liquid culture as determined by persistence of CD34 expression (n=5, p£0.0001) with a delay in CD11b expression (n=5, p£0.05). Initial results by STORM super resolution imaging show loss of LMNA leads to enlarged nuclear size and changes in chromatin localization. In conclusion, loss of LMNA recapitulates features of HSC aging, and restoring LMNA levels may be important for restoring normal function in aged HSCs.

Additional author(s): Emmalee R. Adelman, Dean Wade, Gabriel Gaidosh, Ramin Shiekhattar, Daniel Bilbao, Maria E. Figueroa