Fig. 2. Effects of the Gcm genes in fibroblast cells. (A,B) Staining of mouse Gcm1 (A) and mouse Gcm2 (B) proteins expressed in each stable transformant. The proteins were localized in nuclei. The top right panels show negative controls in which the first antibodies were omitted. (C-F) Gcm gene expression leads to morphological changes in fibroblast cells. Cells were transfected with control (C), mouse Gcm1 (D), mouse Gcm2 (E) or truncated mouse Gcm2 (F) retroviral vectors and cultured for 3 days. Truncated mouse Gcm2 contains the DNA-binding domain but not the transactivating domain. The cells were subsequently stained with X-gal. (G-K) Fibroblast cells were transduced with control (G), mouse Gcm1 (H), mouse Gcm2 (I) or Drosophila gcm (J) viruses at low titers and cultured for 6 days. The cells were stained with X-gal (G-J) and the X-gal⁺ cell number in a cluster was counted. More than 100 colonies for each viral transduction were examined and the distribution of colony size was plotted (K). (L) Total RNA was prepared from stable transformants of control (CT), mouse Gcm1 (a) and mouse Gcm2 (b), and used for RT-PCR analysis of S100ß and ß-actin gene expression. Induction of S100ß expression by mouse Gcm1 and mouse Gcm2 was observed in fibroblast cells. Scale bars: 25 µm in A,B; 50 µm in C-F; 50 µm in G-J.