Knockdown of nucleosome assembly protein 1-like 1 induces mesoderm formation and cardiomyogenesis via notch signaling in murine-induced pluripotent stem cells.

Low efficiency of cardiomyocyte differentiation from induced pluripotent stem cells (iPSCs) hinders the clinical application of iPSC technology for cardiac repair strategy. Recently, we screened out nucleosome assembly protein 1-like 1 (Nap1 vertical bar 1), which was downregulated during the differentiation of P19CL6 cells into cardiomyocytes. Here, we attempted to study the role of Nap1 vertical bar 1 in cardiomyogenesis of iPSC. Nap1 vertical bar 1 was downregulated during the differentiation of iPSC. Knockdown of Nap1 vertical bar 1 dramatically enhanced the differentiation of iPSC into functional cardiomyocytes while overexpression of Nap1 vertical bar 1 sharply lowered the differentiation. Moreover, although Nap1 vertical bar 1 knockdown had little effect on endoderm differentiation, the Nap1 vertical bar 1 modulation significantly accelerated mesoderm development. Re-expressing Nap1 vertical bar 1 in Nap1 vertical bar 1-knockdown-iPSC rescued the effects of Nap1 vertical bar 1. Inducibly overexpressing Nap1 vertical bar 1 at early stage of differentiation greatly inhibited mesoderm induction and cardiogenesis of iPSC. However, mesoderm stem cells (Flk-1-positive cells) originated from Nap1 vertical bar 1-knockdown- or -overexpression-iPSC showed no difference in further cardiomyocyte differentiation compared with that of control-iPSC. Further study revealed that Nap1 vertical bar 1-overexpression increased gamma-secretase activity and the expression of Notch intracellular domain (NICD) and downstream genes during the differentiation of iPSC. gamma-Secretase inhibitor DAPT (N-[ N-(3,5-difluorophenacetyl)-L-alanyl]-S-phenylglycinet-butyl ester) greatly suppressed the production of NICD and abolished the inhibitory effects of Nap1 vertical bar 1-overexpression on mesoderm induction and cardiogenesis. These findings demonstrate that downregulation of Nap1 vertical bar 1 significantly enhances mesodermal induction and subsequent cardiogenesis of murine iPSC via inhibition of gamma-secretase-regulated Notch signaling, which would facilitate the application of iPSC in heart diseases.