In the human, mutations of OTX2 (Orthodenticle homeobox 2 transcription factor) translate into eye malformations of variable expressivity (even between the two eyes of the same individual) and incomplete penetrance, suggesting the existence of subtle thresholds in OTX2 activity. We have addressed this issue by analyzing retinal structure and function in six mutant mice with graded Otx2 activity: Otx2(+/+), Otx2(+/AA), Otx2(+/GFP), Otx2(AA/AA), Otx2(AA/GFP) and Otx2(GFP/GFP). Null mice (Otx2(GFP/GFP)) fail to develop the head and are embryonic lethal, and compound heterozygous Otx2(AA/GFP) mice show a truncated head and die at birth. All other genotypes develop until adulthood. We analyzed eye structure and visual physiology in the genotypes that develop until adulthood and report that phenotype severity parallels Otx2 activity. Otx2(+/AA) are only mildly affected whereas Otx2(+/GFP) are more affected than Otx2(+/AA) but less than Otx2(AA/AA) mice. Otx2(AA/AA) mice later manifest the most severe defects, with variable expressivity. Electrophysiological and histological analyses of the mouse retina revealed progressive death of bipolar cells and cone photoreceptors that is both Otx2 activity- and age-dependent with the same ranking of phenotypic severity. This study demonstrates the importance of gene dosage in the development of age-dependent pathologies and underscores the fact that small gene dosage differences can cause significant pathological states.

Retinal ganglion cell (RGC) axons of binocular animals cross the midline at the optic chiasm (OC) to grow toward their synaptic targets in the contralateral brain. Ventral anterior homeobox 1 (Vax1) plays an essential role in the development of the OC by regulating RGC axon growth in a non-cell autonomous manner. In this study, we identify an unexpected function of Vax1 that is secreted from ventral hypothalamic cells and diffuses to RGC axons, where it promotes axonal growth independent of its transcription factor activity. We demonstrate that Vax1 binds to extracellular sugar groups of the heparan sulfate proteoglycans (HSPGs) located in RGC axons. Both Vax1 binding to HSPGs and subsequent penetration into the axoplasm, where Vax1 activates local protein synthesis, are required for RGC axonal growth. Together, our findings demonstrate that Vax1 possesses a novel RGC axon growth factor activity that is critical for the development of the mammalian binocular visual system.DOI: http://dx.doi.org/10.7554/eLife.02671.001

Mutations of orthodentricle homeobox 2 (OTX2) in human and mice often cause retinal dystrophy and nyctalopia, suggesting a role of OTX2 in mature retina, in addition to its functions in the development of the eye and retina. In support of this, the number of bipolar cells in Otx2 ＋/− post-natal mouse retina was found to be significantly lower than normal. Degeneration of the cells becomes greater as the mice age, leading to the loss of vision. Especially, the type-2 OFF-cone bipolar cells, which do not express Otx2 mRNA but carry Otx2 protein, are most sensitive to Otx2 haplodeficiency. Interestingly, this bipo-lar cell subpopulation imports Otx2 protein from photo-receptors to protect itself from glutamate excitotoxicity in the dark. Moreover, in the bipolar cells, the exogenous Otx2 relocates to the mitochondria to support mitochondrial ATP synthesis. This novel mitochondrial activity of exogenous Otx2 highlights the therapeutic potential of Otx2 protein trans-duction in retinal dystrophy. [BMB Reports 2016; 49(2): 69-70]

RANKL plays an essential role in mammary gland development during pregnancy. However, the molecular mechanism by which RANK signaling leads to mammary gland development is largely unknown. We report here that RANKL stimulation induces phosphorylation of Id2 at serine 5, which leads to nuclear retention of Id2. In lactating Id2Tg; RANKL ؊/؊ mice, Id2 was not phosphorylated and was localized in the cytoplasm. In addition, in lactating Id2 S5A Tg mice, Id2 S5A (with serine 5 mutated to alanine) was exclusively localized in the cytoplasm of mammary epithelial cells (MECs), while endogenous Id2 was localized in the nucleus. Intriguingly, nuclear expression of Id2 S5A rescued increased apoptosis and defective differentiation of MECs in RANKL ؊/؊ mice. Our results demonstrate that nuclear retention of Id2 due to RANK signaling plays a decisive role in the survival and differentiation of MECs during mammary gland development.The mammary gland is a complex organ that proliferates and differentiates during puberty, pregnancy, and lactation under the influence of various hormones, including estrogen, progesterone, and prolactin. At birth, the mammary anlage consists of a few rudimentary ducts that occupy a small portion of the mammary fat pad. Pronounced ductal elongation and side branching commence at puberty. During pregnancy, the complexity of the ductal system increases through the addition of side branches, the formation of lobuloalveolar structures, and the differentiation of secretory epithelia (5,13,14). These proliferation, survival, and lactogenic differentiation steps are essential to form a functional lactating mammary gland during pregnancy.RANKL, a key regulator of osteoclast differentiation and/or activation (1,16,20,22), is essential for the development of a lactating mammary gland during pregnancy (10) and for the expansion of adult mammary stem cells (MaSC) in normal development (2, 17) and breast cancer (11, 36). Mice lacking RANKL or its receptor, RANK, showed impaired lobuloalveolar development during pregnancy, owing to intrinsic defects in the proliferation, survival, and lactogenic differentiation of mammary epithelial cells (MECs). Recent studies have reported that both progesterone (6, 31) and prolactin (40) signaling induce the expression of RANKL, and progesterone induces the expansion of MaSC by a paracrine effecter, RANKL, suggesting that these hormones regulate mammary gland development via the RANKL-RANK signaling pathway (2,11,17,36). However, how RANKL-RANK signaling regulates various aspects of the proliferation, survival, and differentiation of MECs in lactating mammary gland development needs to be elucidated. Cao et al. suggested that activation of a linear RANK-IB kinase ␣ (IKK␣)-NF-B-cyclin D1 signaling cascade could lead to the cellular proliferation of MECs (7). Although we suggested that RANK signaling induces ␤-casein gene expression via CCAAT/enhancer-binding protein ␤ (C/EBP␤) (18), the molecular mechanisms of RANK signaling resulting in the survival and lactogenic differentiat...

The optic neuroepithelial continuum of vertebrate eye develops into three differentially growing compartments: the retina, the ciliary margin (CM), and the retinal pigment epithelium (RPE). Neurofibromin 2 (Nf2) is strongly expressed in slowly expanding RPE and CM compartments, and the loss of mouse Nf2 causes hyperplasia in these compartments, replicating the ocular abnormalities seen in human NF2 patients. The hyperplastic ocular phenotypes were largely suppressed by heterozygous deletion of Yap and Taz, key targets of the Nf2-Hippo signaling pathway. We also found that, in addition to feedback transcriptional regulation of Nf2 by Yap/Taz in the CM, activation of Nf2 expression by Mitf in the RPE and suppression by Sox2 in retinal progenitor cells are necessary for the differential growth of the corresponding cell populations. Together, our findings reveal that Nf2 is a key player that orchestrates the differential growth of optic neuroepithelial compartments during vertebrate eye development.