In vertebrate embryos, the forebrain and eyes are subdivisions of anterior neural tissue. The work here demonstrates a novel regulation of anterior neural development by zebrafish lmo4b. Although these developmental processes have been well studied, it still remains unclear how cells are allotted to and maintained in these subdivided regions. We have identified the zebrafish lmo4b gene and show that it is required to position and maintain proper boundary formation between neural and non-neural ectoderm and by doing so, lmo4b restricts the size of anterior neural tissue. Additionally, lmo4b negatively regulates genes involved in forebrain specification and maintenance such as six3b and rx3, which also promote cell proliferation in anterior neural tissue. Precursors of the peripheral sensory organs (lens, nasal epithelium, inner ear and facial motor neurons) are derived from the placodal primordium that is localized in the boundary region. Therefore, lmo4b may regulate placodal ectoderm development. We also show that lmo4b patterns the eye along the proximodistal axis and is regulated by extracellular signals, Shh and Fgf.
Zebrafish lmo4b is initially expressed in the anterior ectoderm at the neural/non-neural boundary, later in the telencephalon, optic vesicles and finally in optic stalk and retinal pigmented epithelium progenitors. With gain- and loss-of-function studies by overexpression assays and morpholino knockdowns, respectively, we are able to integrate lmo4b into an intricate web of previously identified genes and signaling pathways that regulate embryonic forebrain and eye development. lmo4b overexpression produces embryos with relatively normal gross morphology; however, they have reduced, or no eyes, smaller forebrain and craniofacial and fin defects. The lmo4b knockdown phenotype includes expanded anterior neural plate, followed by enlarged forebrain and eyes, abnormal ears and fins. Moreover, closer analyses reveal that lmo4b morphants have increased mitotic cell and total cell nuclei counts in the optic vesicles.
We propose that because the initial establishment of the presumptive anterior neural boundary is plastic, lmo4b is an essential modulator in positioning the boundary and consequently controls cell commitment to anterior neural fate and tissue growth. We also propose that lmo4b influences the refining of the compartments within the eye through Shh and Fgf regulation.