Incomplete MyoD-induced transdifferentiation is associated with chromatin remodeling deficiencies

Abstract

Our current understanding of cellular transdifferentiation systems is limited. It is oftentimes unknown, at a genome-wide scale, how much transdifferentiated cells differ quantitatively from both the starting cells and the target cells. Focusing on transdifferentiation of primary human skin fibroblasts by forced expression of myogenic transcription factor MyoD, we performed quantitative analyses of gene expression and chromatin accessibility profiles of transdifferentiated cells compared to fibroblasts and myoblasts. In this system, we find that while many of the early muscle marker genes are reprogrammed, global gene expression and accessibility changes are still incomplete when compared to myoblasts. In addition, we find evidence of epigenetic memory in the transdifferentiated cells, with reminiscent features of fibroblasts being visible both in chromatin accessibility and gene expression. Quantitative analyses revealed a continuum of changes in chromatin accessibility induced by MyoD, and a strong correlation between chromatin-remodeling deficiencies and incomplete gene expression reprogramming. Classification analyses identified genetic and epigenetic features that distinguish reprogrammed from non-reprogrammed sites, and suggested ways to potentially improve transdifferentiation efficiency. Our approach for combining gene expression, DNA accessibility, and protein–DNA binding data to quantify and characterize the efficiency of cellular transdifferentiation on a genome-wide scale can be applied to any transdifferentiation system.

The Hox transcription factor Ubx stabilizes lineage commitment by suppressing cellular plasticity in Drosophila

“…The forced expression of TFs expressed only in one lineage like Myoblast Determination Protein (MyoD), which is sufficient to alter cell identity (Tapscott et al, 1988; Weintraub et al, 1989), showed that lineage-restricted TFs can act as major lineage switches by activating lineage-specific transcriptional programs. However, recent profiling experiments revealed that global gene expression and chromatin accessibility changes are imperfect even in MyoD reprogrammed cells, showing that these highly potent reprogramming TFs cannot completely erase the original cell or lineage fate and unequivocally induce the new one (Manandhar et al, 2017). In addition, lineage choice does not only involve the activation of one lineage-specific transcriptional program but also the repression of the programs of all alternative lineages (Kutejova et al, 2016; Mall et al, 2017), a regulatory wiring that needs to be faithfully accomplished in all the different lineages.…”

The Hox Transcription Factor Ubx stabilizes Lineage Commitment by Suppressing Cellular Plasticity

“…Another situation of cell type conversion is cellular reprogramming, which is extensively studied due to its high potential as therapeutic strategy 94 . But although reprogramming strategies have been improved over time, the direct conversion of one somatic cell type into another one, the so-called transdifferentiation, is still inefficient 6,95 . Interestingly, it has been reported recently that the induction of the Hox code typical for a differentiated cell type in pluripotent stem cells (PSCs), either in combination with other factors 96…”

“…We next profiled genome-wide Ubx binding in the same lineages and identical time windows by chromatin immunoprecipitation coupled to massively parallel sequencing (ChIP-seq) using 1x10 6 INTACT-sorted mesodermal and neuronal nuclei and an Ubx specific antibody generated and verified in the lab (see Materials and Methods). The data was benchmarked by the identification of Ubx binding to known target loci.…”

“…The forced expression of TFs expressed only in one lineage like Myoblast Determination Protein (MyoD), which is sufficient to alter cell identity 4,5 , showed that lineage-restricted TFs can act as major lineage switches by activating lineage-specific transcriptional programs. However, recent profiling experiments revealed that global gene expression and chromatin accessibility changes are imperfect even in MyoD reprogrammed cells, showing that these highly potent reprogramming TFs cannot completely erase the original cell or lineage fate and unequivocally induce the new one 6 . In addition, lineage choice does not only involve the activation of one lineage-specific transcriptional program but also the repression of the programs of all alternative lineages 7,8 , a regulatory wiring that needs to be faithfully accomplished in all the different lineages.…”

Poly(ADP-ribose) Polymerase 1 (PARP1) restrains MyoD-dependent gene expression during muscle differentiation

“…Although MyoD can initiate chromatin remodeling and histone modifications at target sites in heterochromatin, nevertheless its binding and transactivation capability are limited by some types of epigenetic constraints. In this regard, the poor ability of MyoD to convert some cell types to the muscle lineage has been ascribed, at least in part, to pre-existing chromatin features that preclude MyoD access to its targets 5 , 21 . For example, it has been reported that trimethylation of lysine 27 on histone H3 (H3K27me3) at the regulatory regions of certain muscle-specific genes prevents MyoD binding and gene activation in undifferentiated myoblasts.…”