Fig. 2. Analysis of the expression of cell cycle regulators in the context of GATA2 loss- and gain-of-function. (A-E) transverse sections of the spinal cord of E10.5 mouse embryos, wild type (A,C,D,E) or Gata2^–/– (B); in situ hybridisation was performed with RNA anti-sense probes for cyclin D1 (A,B,E), cyclin D2 (C) and cyclin D3 (D) genes. In E, further staining, performed with anti-islet 1 antibodies (green), shows that the domain of higher concentration of cyclin D1 transcripts abuts the dorsal limit of motoneurons and overlaps V2 and V1 precursors. Broken lines in A,C-E delineate the presumptive limit between the ventricular and the marginal zones. Brackets in A,B indicate the domain where cyclin D1 transcription is upregulated. (F-H,J-L) Transverse sections of the spinal cord of electroporated chick embryos, hybridised with a chick cyclin D1 (G) or Gata2 (K) antisense RNA probe, then immunostained with anti-HA antibodies (F,J). (H,L) superimposition of E-G (H) and J-K (L). (I,M-T) Confocal analysis of double immunofluorescent staining performed on transverse spinal cord sections of wild-type (I,M-P) or Gata2^–/– (Q-T) E10.5 mouse embryos. Anti-Kip2/p57 antibodies (I,M,Q, green) were coupled with either anti-Isl1 (I, red) or anti-Kip1/p27 (N,R). (O,S) Superimpositions of M,N (O) and Q,R (S). (P,T) Double immunostaining with anti-p27 and anti-Isl1 antibodies, respectively visualised with anti-mouse IgG1 coupled to Alexa Fluor 546 and anti mouse IgG2b coupled to Alexa Fluor 488. The respective presumptive domains of V1, V2 and motor (MN) neurons are indicated in I and P. White arrows in I indicate that the V2 presumptive domain does not express p57/Kip2, and in R,S,T, that the same domain lacks p27 expression in Gata2^–/– embryos. Scale bar: 70 µm in A-D; 60 µm in E-L; 80 µm in M-T.