Postmitotic accumulation of histone variant H3.3 in new cortical neurons establishes neuronal chromatin, transcriptome, and identity

Abstract

Histone variants, which can be expressed outside of S-phase and deposited DNA synthesis-independently, provide long-term histone replacement in postmitotic cells, including neurons. Beyond replenishment, histone variants also play active roles in gene regulation by modulating chromatin states or enabling nucleosome turnover. Here, we uncover crucial roles for the histone H3 variant H3.3 in neuronal development. We find that newborn cortical excitatory neurons, which have only just completed replication-coupled deposition of canonical H3.1 and H3.2, substantially accumulate H3.3 immediately postmitosis. Codeletion of H3.3-encoding genes H3f3a and H3f3b from newly postmitotic neurons abrogates H3.3 accumulation, markedly alters the histone posttranslational modification landscape, and causes widespread disruptions to the establishment of the neuronal transcriptome. These changes coincide with developmental phenotypes in neuronal identities and axon projections. Thus, preexisting, replication-dependent histones are insufficient for establishing neuronal chromatin and transcriptome; de novo H3.3 is required. Stage-dependent deletion of H3f3a and H3f3b from 1) cycling neural progenitor cells, 2) neurons immediately postmitosis, or 3) several days later, reveals the first postmitotic days to be a critical window for de novo H3.3. After H3.3 accumulation within this developmental window, codeletion of H3f3a and H3f3b does not lead to immediate H3.3 loss, but causes progressive H3.3 depletion over several months without widespread transcriptional disruptions or cellular phenotypes. Our study thus uncovers key developmental roles for de novo H3.3 in establishing neuronal chromatin, transcriptome, identity, and connectivity immediately postmitosis that are distinct from its role in maintaining total histone H3 levels over the neuronal lifespan.

A Vector System Encoding Histone H3 Mutants Facilitates Manipulations of the Neuronal Epigenome

“…This observation might reflect the fact that neuronal differentiation is accompanied by global changes in chromatin modifications 54 . Supporting this idea, recent findings indicate that accumulation of the histone H3.3 isoform in differentiating cortical neurons is important to establish their identity and axonal connectivity 55 . Therefore, the induction and incorporation into nucleosomes of histone H3.3 during cortical neurogenesis could favor the generation of methylation-depleted histones in the presence of K-to-M mutants.…”

Knockout tales: the versatile roles of histone H3.3 in development and disease

“…A recent paper studied the effect of H3f3a and H3f3b co-deletion specifically in developing neurons [ 19 ]. They knocked out H3f3a and H3f3b in neural progenitor cells using Emx1-Cre, and specifically in excitatory neurons after terminal mitosis using Neuro6d-Cre.…”

“…They knocked out H3f3a and H3f3b in neural progenitor cells using Emx1-Cre, and specifically in excitatory neurons after terminal mitosis using Neuro6d-Cre. In both situations, knockout mice were born alive but died within a few hours of birth [ 19 ]. These mice show changes to neuronal fate and identity as well as defects in axon projection and development.…”

“…These mice show changes to neuronal fate and identity as well as defects in axon projection and development. Transcriptomic and epigenomic analyses found that accumulation of H3.3 in chromatin is required for properly setting up the transcriptome in newly post-mitotic neurons and regulating the levels of H3K4me3 and H3K27me3 at these genes [ 19 ].…”

Neuronal cell cycle reentry events in the aging brain are more prevalent in neurodegeneration and lead to cellular senescence

“…Terminally differentiated neurons are believed to have irreversibly exited from the cell division process [ 1 ], and by various means, these cells obtain their postmitotic identity [ 2 , 3 ]. Nonetheless, a large body of work has indicated that as individuals age, a small population of these cells may recommit to a cell cycle-like process [ 4 ], and increasing levels of these events are found in disease-affected brains of patients with late-onset Alzheimer’s disease (AD) [ 4 – 11 ].…”