Fig. 1. Necab encodes a phylogenetically conserved mammalian protein. (A) Nucleotide sequence and amino acid translation of mouse Necab cDNA. Necab encodes a 389 amino acid protein (dark blue) with an EF-hand Ca²⁺-binding domain (in pale blue). A polyadenylation signal is present at position 1944 to 1953 of the nucleotide sequence. (B) Alignment of mouse (Mus musculus; Accession Number AF411253), macacus (Macaca fascicularis; Accession Number AB046031) and human (Homo sapiens) Necab amino acid sequence. Red, amino acid identity in all shown sequences; blue, amino acid identity in two out of three species; green, amino acid similarity; black, divergence. The amino acid stretch in bold of the human sequence represents possible mistakes in the predicted translation product of NECAB cDNA. (C, part I) Immunohistochemistry with Ab1 antibody at 10.5 dpc (coronal sections) showing immunoreactivity in the retinal-pigmented epithelium (rpe) and in the neural retina (nr). No signal is detected in the in the lens (ls) or in the surrounding eye mesenchyme. (C, part II) Immunoreactivity of Ab1 in the Rathke’s pouch (Rp) and in the infundibulum recess (inf). (C, part III) Immunofluorescence at 17.5 dpc (cross sections) with Ab1 antibody. Predominant immunoreactivity is found in the cytoplasm of the ganglion cells (arrowheads). (C, part IV) A Necab/v5 epitope fusion protein is localized to the cytoplasm in COS cells. (D) FISH on metaphase chromosomes. NECAB is localized to human chromosome 16q23 and Necab is localized to mouse chromosome 8E1.