Under natural conditions, tissue and cellular differentiation along defined lineages is characterized by an inexorably forward-moving process that terminates in highly specialized cells. Waddington, following Morgan (Morgan, 1901
), characterized the process of development as essentially “epigenetic” (from “epigenesis”) (Waddington, 1939
) and also introduced the metaphor of an “epigenetic landscape” (Waddington, 1940
), which he depicted with a ball rolling down a hill of bifurcating valleys symbolizing the specification of defined cell lineages and fates during the progress of differentiation (Waddington, 1939
). It is notable that Waddington's usage of epigenetic to denote the origination and propagation of information about cellular states during differentiation differs considerably from its recent reformulation to mean “on the genome” and its association with chemical modifications to DNA or chromatin (Ptashne, 2007
). Here we employ the classical usage throughout.
Waddington astutely reasoned that epigenesis is a “historical” process requiring a memory “faculty” to keep directed lineage programs on track (Waddington, 1939
). Indeed, developing cells are frequently exposed to stimuli, whether exogenous (e.g., a morphogen) or endogenous (e.g., a transcription factor [TF]), that can permanently alter cellular fate. Whether or in what form cells in fact maintain information concerning prior developmental fate decisions during epigenesis is currently unknown.
The epigenetic landscape paradigm has also been invoked to explain abnormal processes such as oncogenesis (Pujadas and Feinberg, 2012
). Cancer cells are widely described as being “de-differentiated” compared with their normal counterparts, based on limited analyses of metabolic (Warburg, 1956
), histological (Gleason and Mellinger, 1974
), gene-activity (Hirszfeld et al., 1932
; Tatarinov, 1964
), and proliferative and self-renewal phenotypes (Beard, 1902
; Waddington, 1935
). However, quantifying this concept and generalizing it beyond a few selected markers have proven difficult.
Chromatin structure represents a highly plastic vehicle for specifying cellular regulatory states and is a conceptually attractive template for recording and transmitting epigenetic information (Bernstein et al., 2006
; Hawkins et al., 2010
; Paige et al., 2012
; Wamstad et al., 2012
; Zhu et al., 2013
). DNase I-hypersensitive sites (DHSs) represent focal alterations in the primary structure of chromatin that result from engagement of sequence-specific transcription factors in place of a canonical nucleosome (Gross and Garrard, 1988
; Thurman et al., 2012
). In a classic experiment, Groudine and Weintraub demonstrated that induced DHSs could be propagated to, and stably perpetuated by, daughter cells even after the inducing stimulus had been withdrawn (Groudine and Weintraub, 1982
). This result suggests that newly arising DHSs created by TF occupancy of quiescent regulatory DNA have the potential to encode cellular states and to perpetuate that information through continued TF occupancy in daughter cells. Whether, or to what extent, such a mechanism operates during normal development and differentiation, however, is currently unknown.
To explore the role of TF-driven chromatin structure at regulatory DNA in normal and transformed cells during epigenesis, we analyzed genome-wide patterns of DHSs across a wide array of cell types and states, including definitive adult primary cells, embryonic stem cells (ESCs), cells undergoing directed lineage differentiation from ESCs to cardiomyocytes, and diverse cancer cell types. Our findings, detailed below, are interpreted to indicate four fundamental conclusions. First, patterns of DHSs in definitive cells encode “memory” of early developmental fate decisions that establish lineage hierarchies. Second, lineage differentiation couples the extensive activation of novel regulatory DNA compartments with propagation and sequential restriction of the ES DHS landscape as a function of cellular maturity. Third, developmentally stable DHSs chiefly encode binding sites for self-regulating TFs, suggesting a mechanistic role for TF-encoded feedback circuits in propagating developmental information. Finally, oncogenesis is accompanied by a disordered retrograde remodeling of the regulatory DNA landscape in a fashion that defies normal developmental pathways and departs fundamentally from the paradigm of the epigenetic landscape. Together these findings indicate a central role for patterning and propagation of regulatory DNA marked by DHSs in the genesis and proper maintenance of developmental programs.