An RNAi Screen Identifies Msi2 and Prox1 as Having Opposite Roles in the Regulation of Hematopoietic Stem Cell Activity

In this study, we describe an in vivo RNA interference functional genetics approach to evaluate the role of 20 different conserved polarity factors and fate determinants in mouse hematopoietic stem cell (HSC) activity. In total, this screen revealed three enhancers and one suppressor of HSC-derived reconstitution. Pard6a, Prkcz, and Msi2 shRNA-mediated depletion significantly impaired HSC repopulation. An in vitro promotion of differentiation was observed after the silencing of these genes, consistent with their function in regulating HSC self-renewal. Conversely, Prox1 knockdown led to in vivo accumulation of primitive and differentiated cells. HSC activity was also enhanced in vitro when Prox1 levels were experimentally reduced, identifying it as a potential antagonist of self-renewal. HSC engineered to overexpress Msi2 or Prox1 showed the reverse phenotype to those transduced with corresponding shRNA vectors. Gene expression profiling studies identified a number of known HSC and cell cycle regulators as potential downstream targets to Msi2 and Prox1. [Display omitted] ► RNAi screening examines roles of conserved polarity factors in HSC regulation ► Screen identified three HSC agonists (Msi2, Pard6a, Prkcz) and one antagonist (Prox1) ► Overexpression and knockdown of Msi2 and Prox1 gave reciprocal phenotypes ► Msi2 and Prox1 modulate the expression of self-renewal and cell cycle regulators