p73 regulates maintenance of neural stem cell

Abstract

p73, a member of the p53 family, is a transcription factor that plays a key role in many biological processes. In the present study, we show that TAp73 is expressed in neural stem cells (NSC) and its expression increases following their differentiation. NSC from p73 null mice have a reduced proliferative potential, together with reduced expression of members of the Sox-2 and Notch gene families known to be important for NSC proliferation. In parallel with this in vitro data, the width of the neurogenic areas was reduced in the brains of embryonic and adult p73-/- mice. These data suggest that p73, and in particular TAp73, is important for maintenance of the NSC pool.

Fig. 1

Expression of p53 family members in NSC. (A) NSC were cultured as described in Section 2. Cells were harvested at the indicated time points and expression of TAp73, ΔNp73 and p53 was evaluated by qPCR. Data are normalized to the housekeeping gene Gapdh and are relative to the expression of TAp73. Data represent mean ± SD of three different experiments. (B and C) An NSC single cell suspension was allowed to differentiate in vitro and expression of the indicated genes was evaluated by qPCR. Data are normalized to the housekeeping gene Gapdh relative to DIV 1. A representative experiment of three independent experiments is shown.

Fig. 2

p73-deficient NSC show decreased proliferation. (A) E14.5 neurospheres were derived from wild-type and p73−/− mice. A representative (two different fields per each genotyping) picture of NSC is shown. (B) Diameter of NSC from p73+/+ and p73−/− mice at DIV 4 and 7, respectively. (C) Size distribution of NSC at DIV 7. p73−/− NSC accumulate in the lower range. Data represent mean ± SD of three different experiments.

Fig. 3

Loss of p73 impairs expression of genes involved in NSC self-renewal. Relative expression of the indicated genes in neurospheres derived from p73−/− mice at DIV 7. Data are normalized to the housekeeping gene Gapdh and relative to their expression in p73+/+ NSC. Data represent mean ± SD of three different experiments.

Fig. 4

Reduction of neurogenic areas in p73−/− mice. (A) Nestin immunofluorescence of the ventricular zone in coronal sections in E17.5 mice. Magnification 20×. Graph shows the mean ± SD of thickness of neurogenic area. (B) The dentate gyrus from P7 p73+/+ and p73−/− mice was stained with antibodies to GFAP and nestin. Arrows indicate double positive cells. Magnification 40×. Graph shows the mean ± SD of GFAP/nestin positive cells. VZ = ventricular zone; GCL = granular cell layer; Hil = hilus.