Class II HDACs are required during early stage mouse embryonic stem cell differentiation in response to Nitric Oxide.

INTRODUCTION & BACKGROUND: We have recently described that in mouse embryonic stem cells (ES) laminar shear stress provides chromatin programming signals. This effect is similar to that of nitric oxide (NO) which directly promotes cardiomyogenesis underlying the possible presence of a common mechanism of action the characterization of which has been the object of the present study. METHODS & RESULTS: ES were treated with the NO donor DETANO from 1 to 24 hours (h) in the presence of absence of leukaemia inhibitory factor (LIF). In LIF-deprived cells the NO donor determined a marked global reduction in histone H3 acetylation paralleled by the expression of a number of cardiovascular markers. In this condition the stemness-associated gene product Oct4 decreased rapidly at mRNA and protein level underlying the potential opening of a NO-dependent differentiation program. Further class-II histone deacetylase (HDAC) 4, 5, and 7, rapidly shuttled to nucleus upon LIF deprivation. This phenomenon was transient as HDAC molecules returned to the cytoplasm in about 6 h. Intriguingly, the exposure of ES to the NO donor allowed the nuclear retention of these enzymes still detectable at the 6 h time-point. Further, we found that NO treatment stimulated the formation of nuclear complexes including the class II HDACs 4, 5 and the protein phosphatase 2A. Remarkably, ES treatment with the class II selective inhibitor MC 1568 prevented ES differentiation in the presence of NO. CONCLUSIONS: These results suggest a potential role for class II HDACs in the NO-dependent early stage of ES commitment upon LIF removal.