Depletion of human histone H1 variants uncovers specific roles in gene expression and cell growth.
Bottom Line: Microarray experiments have shown a different subset of genes to be altered in each H1 knock-down.Interestingly, H1.2 depletion caused specific effects such as a cell cycle G1-phase arrest, the repressed expression of a number of cell cycle genes, and decreased global nucleosome spacing.On its side, H1.4 depletion caused cell death in T47D cells, providing the first evidence of the essential role of an H1 variant for survival in a human cell type.
Affiliation: Centre de Regulació Genòmica, Barcelona, Spain.
At least six histone H1 variants exist in somatic mammalian cells that bind to the linker DNA and stabilize the nucleosome particle contributing to higher order chromatin compaction. In addition, H1 seems to be actively involved in the regulation of gene expression. However, it is not well known whether the different variants have distinct roles or if they regulate specific promoters. We have explored this by inducible shRNA-mediated knock-down of each of the H1 variants in a human breast cancer cell line. Rapid inhibition of each H1 variant was not compensated for by changes of expression of other variants. Microarray experiments have shown a different subset of genes to be altered in each H1 knock-down. Interestingly, H1.2 depletion caused specific effects such as a cell cycle G1-phase arrest, the repressed expression of a number of cell cycle genes, and decreased global nucleosome spacing. On its side, H1.4 depletion caused cell death in T47D cells, providing the first evidence of the essential role of an H1 variant for survival in a human cell type. Thus, specific phenotypes are observed in breast cancer cells depleted of individual histone H1 variants, supporting the theory that distinct roles exist for the linker histone variants.
Related in: MedlinePlus
Mentions: Upon Dox treatment, we observed differences in growth rate among H1 variant knock-downs, in particular, H1.2 and H1.4 depleted cells failed to reach confluency (Figure 2A) and exhibited a slower growth rate. To quantify this we mixed, at a 1∶1 ratio, each of the shRNA expressing cell lines (RedFP and GFP-positive upon Dox treatment) with parental T47D cells (RedFP and GFP-negative), and we monitored by FACS the proportion between the two populations over time in culture in the presence of Dox (Figure 2B). Slow progression of H1.4 knocked-down cells was seen at six days after Dox addition and of H1.2 depleted cells at day 9. At day 12, H1.5 depleted cells were also less abundant than the parental ones. In addition, at day 6, H1.4 cells had changed morphology towards a necrotic phenotype (Figure 2A and Figure S2).