Fig. 1.

Mice deficient for NONO show increased cell division and reduced senescence. (A) NONO RNA expression measured by qPCR in various tissues taken from WT (+) and Nono^gt animals (gt, not detectable). y axis, expression levels relative to maximum observed expression. Br, brain; De, dermis; He, heart; Ki, kidney; Li, liver; Lu, lung (n = 4 animals). (Inset) NONO protein measured in liver nuclear extract pooled from two representative animals, as well as in unrelated C57-Bl6J mice (Bl6/J). (B) WT and Nono^gt primary fibroblasts were counted and passaged every 2 d and a constant number of cells plated to a new dish. Total cell number over time is plotted relative to initial cell number as population doublings. (Student t test for significant difference of doubling rates, P = 0.05.) (C) Cells from each passage in B were stained for SA-βgal activity, and the percentages of total cells expressing this marker were recorded. In this and all subsequent figures, *P < 0.05 and **P < 0.01 (Student t test). (D) Duplicate nonconfluent plates of WT and Nono^gt primary fibroblasts were stained with CFSE and allowed to divide for 4 additional days. Dye intensity was then measured by flow cytometry (duplicate plates of cells in green and blue). As a control, other plates of the same cells were treated with mitomycin C to inhibit cell division immediately after staining and then treated in parallel (red). Numbers near curves reflect the percentage overlap between the green/blue curves and red curve. (E) WT and Nono^gt primary fibroblasts were infected with lentivirus WPI (expressing GFP) or WPI-NONO (expressing NONO) and allowed to proliferate via serial passaging as in A. Relative cell number for each cell type ±SD was plotted at each passage (i.e., every 2 d).