Fig. 3

Rescue ability of invertebrate SoxB homologs. a Amino acid alignments of the HMG-box of mouse GroupB1 family members and SoxB homologs from amphioxus, ascidian (Ci-SoxB1) and fluitfly (SoxN). Non-homologous amino acids are highlighted in red, the 3 helices of the DNA binding domain are indicated as yellow bars. b Cartoon of SoxB protein structures. All share a very similar 79 amino acid HMG-box but the N- and C-terminal regions vary in their length and degree of homology to Sox2 (shown as %). c A maximum likelihood phylogenetic tree of SoxB homologs. d Reporter gene assays with SoxB homologs. Relative activities are indicated as ‘fold activation’ on the log scale. Each assay represents the mean of 3 replicates, error bars mean standard deviations. All factors show comparable ability to activate the Sox2 reporter. e ES cell rescue assays. The numbers of primary colonies and standard deviation obtained with each GroupB Sox protein are indicated under the ID Colums represent: 1 – primary transfectants in the absence of Tc. 2 – Secondary colonies in the presence of Tc. 3 – Colonies at passage 3 in the presence of Tc. All Sox factors assayed showed rescue ability. f Immunostaining of the rescued ES cells. Sox2 (poly) indicates staining with goat anti-Sox2 antibody which shows weak cross-reactivity with SoxB homologs, except for the fly SoxN protein. Sox2 (mono) indicates staining with a mouse anti-Sox2 monoclonal antibody specifically reactive with mouse Sox2, confirming the proper loss of mouse Sox2 in the rescued ES cells