In the zebrafish brain, strong GbX-staining was found in the oculomotor nucleus and in nerves with axons innervating muscles that control the movements of the eye. The n. oculomotorius exits the brain ventrally and passes the hypothalamus, which may explain the immunostaining in this region. In sagittal sections the hypothalamic corpus mamillare and fasciculus retroflexus exhibit prominent staining as well. The mamillary body is a pair of nuclei that receives and relays olfactory impulses. The f. retroflexus is a fiber tract that connects the habenula with the midbrain and hindbrain. The vertebrate retina is composed of three layers of nerve cell bodies and two layers of synapses. GbX is localized in the ganglion cell layer of the retina, which contains the nuclei of the ganglion cells and some displaced amacrine cells. The ganglion cells project visual signals from the photoreceptors to the tectum opticum, which represents the major visual center in teleosts. In summary, GbX appears to be associated mainly with neurons of the sensory system. N-terminal lipid attachment results in association of the acylated protein with the cytoplasmic side of the membrane. Our results suggest that GbX is indeed myristoylated at Gly2 and palmitoylated at Cys3. Myristoylation is a covalent and irreversible attachment of the fatty acid myristate, which is cotranslationally catalyzed by N-myristoyltransferase. This enzyme recognizes an N-terminal Gly, which is exposed by removal of the initiator Met. Unlike myristoylation, palmitoylation is reversible and therefore plays a role in regulatory functions, subcellular trafficking and localization. Palmitate is posttranslationally attached to the protein by multiple enzymes. Myristoylation of GbX is GDC-0879 essential for membrane localization, whereas palmitoylation is required for full association. A small proportion of GbX protein appears to be palmitoylated and localized at the membrane even in the absence of a prior myristoylation. The complete lack of acylation, as in the GbX-mutant constructs, resulted in an accumulation of the GFP-tagged protein in the nucleus. Our results thus indicate that both lipid modifications are necessary for correct subcellular localisation of GbX. Hb, Mb, and Ngb of vertebrates are proteins located in the cytoplasm. Cygb is a cytoplasmic protein in fibroblasts and related cells, but partly resides in the nucleus of some neurons. Membrane-bound globins had been unknown in vertebrates, but have previously been reported in bacteria and such a protein has also been identified in the gills of the green shore crab, Carcinus maenas. Although the crab globin harbors an Nterminal N-myristoylation site, it is evolutionary not related to GbX, suggesting convergent evolution of membrane attachment in eukaryotes. To the best of our knowledge, GbX is therefore the first example in vertebrates where a globin is attached to the membrane. Nevertheless, membrane association of globins may be more widespread in animals than currently acknowledged and hint to a common but still poorly defined function of globins.