In the developing retina, neurogenesis and cell differentiation are coupled with cell proliferation. However, molecular mechanisms that coordinate cell proliferation and differentiation are not fully understood. In this study, we found that retinal neurogenesis is severely delayed in the zebrafish stem-loop binding protein (slbp) mutant. SLBP binds to a stem-loop structure at the 3'-end of histone mRNAs, and regulates a replication-dependent synthesis and degradation of histone proteins. Retinal cell proliferation becomes slower in the slbp1 mutant, resulting in cessation of retinal stem cell proliferation. Although retinal stem cells cease proliferation by 2 days postfertilization (dpf) in the slbp mutant, retinal progenitor cells in the central retina continue to proliferate and generate neurons until at least 5dpf. We found that this progenitor proliferation depends on Notch signaling, suggesting that Notch signaling maintains retinal progenitor proliferation when faced with reduced SLBP activity. Thus, SLBP is required for retinal stem cell maintenance. SLBP and Notch signaling are required for retinal progenitor cell proliferation and subsequent neurogenesis. We also show that SLBP1 is required for intraretinal axon pathfinding, probably through morphogenesis of the optic stalk, which expresses attractant cues. Taken together, these data indicate important roles of SLBP in retinal development.

Genetic mutations in aryl hydrocarbon receptor interacting protein-like 1 (AIPL1) cause photoreceptor degeneration associated with Leber congenital amaurosis 4 (LCA4) in human patients. Here we report retinal phenotypes of a zebrafish aipl1 mutant, gold rush (gosh). In zebrafish, there are two aipl1 genes, aipl1a and aipl1b, which are expressed mainly in rods and cones, respectively. The gosh mutant gene encodes cone-specific aipl1, aipl1b. Cone photoreceptors undergo progressive degeneration in the gosh mutant, indicating that aipl1b is required for cone survival. Furthermore, the cone-specific subunit of cGMP phosphodiesterase 6 (Pde6c) is markedly decreased in the gosh mutant, and the gosh mutation genetically interacts with zebrafish pde6c mutation eclipse (els). These data suggest that Aipl1 is required for Pde6c stability and function. In addition to Pde6c, we found that zebrafish cone-specific guanylate cyclase, zGc3, is also decreased in the gosh and els mutants. Furthermore, zGc3 knockdown embryos showed a marked reduction in Pde6c. These observations illustrate the interdependence of cGMP metabolism regulators between Aipl1, Pde6c, and Gc3 in photoreceptors.

Many abiotic stresses induce the generation of nitric oxide (NO) in plant tissues, where it functions as a signal molecule in stress responses. Plants modulate NO by oxidizing it to NO 3 − with plant hemoglobin (GLB), because excess NO is toxic to cells. At least eight genes encoding GLB have been identified in soybean, in three clades: GLB1, GLB2, and GLB3. However, it is still unclear which GLB genes are responsible for NO regulation under abiotic stress in soybean. We exposed soybean roots to flooding, salt, and two NO donors-sodium pentacyanonitrosylferrate (III) dihydrate (SNP) and S-nitroso-N-acetyl-d,l-penicillamine (SNAP)-and analyzed expression of GLB genes. GmGLB1, one of two GLB1 genes of soybean, significantly responded to both SNP and SNAP, and its induction was almost completely repressed by a NO scavenger, 2-(4-carboxyphenyl)-4,4,5,5-tetramethylimidazoline-1-oxyl-3-oxide. GmGLB1 responded to flooding but not to salt, suggesting that it is responsible for NO regulation under NO-inducing abiotic stresses such as flooding. GmGLB3, one of two GLB3 genes of soybean, did not respond to NO donors at all but did respond to flooding, at a lower level than GmGLB1. These results suggest that flooding induces not only NO but also unknown factor(s) that induce GmGLB3 gene in soybean.

Phytohormone abscisic acid (ABA) inhibits root nodule formation of leguminous plants. LjGlu1, a β-1,3-glucanase gene of Lotus japonicus, has been identified as an ABA responsive gene. RNA interference of LjGlu1 increased nodule number. This suggests that LjGlu1 is involved in the regulation of nodule formation. Host legumes control nodule number by autoregulation of nodulation (AON), in which the presence of existing root nodules inhibits further nodulation. For further characterization of LjGlu1, we focused on the expression of LjGlu1 in relation to AON. In a split-root system, LjGlu1 expression peaked when AON was fully induced. Hairy roots transformed with LjCLE-RS1, a gene that induces AON, were generated. Expression of LjGlu1 was greater in the transgenic roots than in untransformed roots. LjGlu1 was not induced in a hypernodulating mutant inoculated with Mesorhizobium loti. These results suggest that the expression of LjGlu1 is involved in the system of AON. However, neither hypernodulation nor enlarged nodulation zone was observed on the transgenic hairy roots carrying LjGlu1-RNAi, suggesting that LjGlu1 is not a key player of AON. Recombinant LjGlu1 showed endo-β-1,3-glucanase activity. LjGlu1-mOrange fusion protein suggested that LjGlu1 associated with M. loti on the root hairs. Exogenous β-1,3-glucanase inhibited infection thread formation by both the wild type and the mutant, and nodule numbers were reduced. These results suggest that LjGlu1 is expressed in response to M. loti infection and functions outside root tissues, resulting in the inhibition of infection.