β-actin regulates a heterochromatin landscape essential for optimal induction of neuronal programs during direct reprograming

Abstract

During neuronal development, β-actin serves an important role in growth cone mediated axon guidance. Consistent with this notion, in vivo ablation of the β-actin gene leads to abnormalities in the nervous system. However, whether β-actin is involved in the regulation of neuronal gene programs is not known. In this study, we directly reprogramed β-actin+/+ WT, β-actin+/- HET and β-actin-/- KO mouse embryonic fibroblast (MEFs) into chemically induced neurons (CiNeurons). Using RNA-seq analysis, we profiled the transcriptome changes among the CiNeurons. We discovered that induction of neuronal gene programs was impaired in KO CiNeurons in comparison to WT ones, whereas HET CiNeurons showed an intermediate levels of induction. ChIP-seq analysis of heterochromatin markers demonstrated that the impaired expression of neuronal gene programs correlated with the elevated H3K9 and H3K27 methylation levels at gene loci in β-actin deficient MEFs, which is linked to the loss of chromatin association of the BAF complex ATPase subunit Brg1. Together, our study shows that heterochromatin alteration in β-actin null MEFs impedes the induction of neuronal gene programs during direct reprograming. These findings are in line with the notion that H3K9Me3-based heterochromatin forms a major epigenetic barrier during cell fate change.

β-actin dependent chromatin remodeling mediates compartment level changes in 3D genome architecture

“…A potential relationship between β-actin and BAF/PRC mediated chromatin remodeling raises the possibility that nuclear β-actin levels may play a regulatory role in shaping the chromatin landscape during development. Such a function is consistent with the wellestablished roles of actin in transcriptional reprogramming (16) and our previous work showing that βactin knockout cells exhibit specific defects in gene regulation during neural reprogramming (7). We therefore hypothesize that regulation of nuclear β-actin levels may potentially influence both local chromatin accessibility and higher-order genome organization.…”

Emerging roles of cytoskeletal proteins in regulating gene expression and genome organization during differentiation

“…Recent work has shown that the nuclear β-actin pool significantly contributes to neuronal development by regulating genome organization and activation of neuronal programs during neurogenesis. To address this question, we directly reprogrammed wild-type and β-actin knockout embryonic fibroblasts as well as embryonic fibroblasts from a heterozygous mouse where only one of the βactin alleles was disrupted [17]. Compromising βactin levels was associated with impaired expression of neural and proneural genes involved in the neuronal cytoskeleton, synapse, axon development and guidance, voltage-gated ion channels and neuronrelated transcription factors.…”

“…Compromising βactin levels was associated with impaired expression of neural and proneural genes involved in the neuronal cytoskeleton, synapse, axon development and guidance, voltage-gated ion channels and neuronrelated transcription factors. Notably, members of the Zic and Irx family of genes, which are involved in the onset of neuronal fate [93], were severely down-regulated in the absence of β-actin [17]. The overall impaired induction of neuronal genes in βactin knockouts was correlated with increased H3K9Me3 levels and loss of Brg1 (ATPase subunit of BAF chromatin remodeling complex) occupancy at transcription start sites of multiple gene loci, including Zic and Irx genes.…”

“…Furthermore, HP1α protein is known to homodimerize and bridge H3K9 methylated nucleosomes while PRC1 proteins are thought to induce clustering of polycomb bound chromatin and nucleosomes into phase separated 'polycomb bodies' implicated in the regulation of genome architecture [105] and maintenance of long-range chromatin interactions [106]. β-actin depletion can potentially affect the regulation of both HP1α and PRC1 proteins [17,72]. We have shown that β-actin knockout cells exhibit redistribution of HP1αpositive heterochromatin and loss of chromatin association of Brg1, the essential ATPase subunit of the BAF complex [72].…”

“…Furthermore, actin affects chromatin organization by associating with or being a component of developmentally important chromatin remodelers such as INO80 and TIP60 complexes in yeast and BAF (Brahma-associated factors) complex in mammals [1]. As the BAF complex has important roles in lineage specification and cell fate reprogramming, actin depletion has also been shown to affect the induction of differentiation programs in cells undergoing transcriptional reprogramming [17]. Emerging evidence also shows that mechanical force can affect actin dynamics and chromatin regulation [18].…”