Genetics of photoreceptor degeneration and regeneration in zebrafish

Zebrafish are unique in that they provide a useful model system for studying two critically important problems in retinal neurobiology, the mechanisms responsible for triggering photoreceptor cell death and the innate stem cell–mediated regenerative response elicited by this death. In this review we highlight recent seminal findings in these two fields. We first focus on zebrafish as a model for studying photoreceptor degeneration. We summarize the genes currently known to cause photoreceptor degeneration, and we describe the phenotype of a few zebrafish mutants in detail, highlighting the usefulness of this model for studying this process. In the second section, we discuss the several different experimental paradigms that are available to study regeneration in the teleost retina. A model outlining the sequence of gene expression starting from the dedifferentiation of Müller glia to the formation of rod and cone precursors is presented

The Bacterial Fimbrial Tip Acts as a Mechanical Force Sensor

The subunits that constitute the bacterial adhesive complex located at the tip of the fimbria form a hook-chain that acts as a rapid force-sensitive anchor at high flow

Structural Basis for Type VI Secretion Effector Recognition by a Cognate Immunity Protein

The type VI secretion system (T6SS) has emerged as an important mediator of interbacterial interactions. A T6SS from Pseudomonas aeruginosa targets at least three effector proteins, type VI secretion exported 1–3 (Tse1–3), to recipient Gram-negative cells. The Tse2 protein is a cytoplasmic effector that acts as a potent inhibitor of target cell proliferation, thus providing a pronounced fitness advantage for P. aeruginosa donor cells. P. aeruginosa utilizes a dedicated immunity protein, type VI secretion immunity 2 (Tsi2), to protect against endogenous and intercellularly-transferred Tse2. Here we show that Tse2 delivered by the T6SS efficiently induces quiescence, not death, within recipient cells. We demonstrate that despite direct interaction of Tsi2 and Tse2 in the cytoplasm, Tsi2 is dispensable for targeting the toxin to the secretory apparatus. To gain insights into the molecular basis of Tse2 immunity, we solved the 1.00 Å X-ray crystal structure of Tsi2. The structure shows that Tsi2 assembles as a dimer that does not resemble previously characterized immunity or antitoxin proteins. A genetic screen for Tsi2 mutants deficient in Tse2 interaction revealed an acidic patch distal to the Tsi2 homodimer interface that mediates toxin interaction and immunity. Consistent with this finding, we observed that destabilization of the Tsi2 dimer does not impact Tse2 interaction. The molecular insights into Tsi2 structure and function garnered from this study shed light on the mechanisms of T6 effector secretion, and indicate that the Tse2–Tsi2 effector–immunity pair has features distinguishing it from previously characterized toxin–immunity and toxin–antitoxin systems

In the Absence of Sonic Hedgehog, p53 Induces Apoptosis and Inhibits Retinal Cell Proliferation, Cell-Cycle Exit and Differentiation in Zebrafish

Background: Sonic hedgehog (Shh) signaling regulates cell proliferation during vertebrate development via induction of cell-cycle regulator gene expression or activation of other signalling pathways, prevents cell death by an as yet unclear mechanism and is required for differentiation of retinal cell types. Thus, an unsolved question is how the same signalling molecule can regulate such distinct cell processes as proliferation, cell survival and differentiation. Methodology/Principal Findings: Analysis of the zebrafish shh 2/2 mutant revealed that in this context p53 mediates elevated apoptosis during nervous system and retina development and interferes with retinal proliferation and differentiation. While in shh 2/2 mutants there is activation of p53 target genes and p53-mediated apoptosis, an increase in Hedgehog (Hh) signalling by over-expression of dominant-negative Protein Kinase A strongly decreased p53 target gene expression and apoptosis levels in shh 2/2 mutants. Using a novel p53 reporter transgene, I confirm that p53 is active in tissues that require Shh for cell survival. Proliferation assays revealed that loss of p53 can rescue normal cell-cycle exit and the mitotic indices in the shh 2/2 mutant retina at 24, 36 and 48 hpf. Moreover, generation of amacrine cells and photoreceptors was strongly enhanced in the double p53 2/2 shh 2/2 mutant retina suggesting the effect of p53 on retinal differentiation. Conclusions: Loss of Shh signalling leads to the p53-dependent apoptosis in the developing nervous system and retina

Phenylthiourea Specifically Reduces Zebrafish Eye Size

Phenylthiourea (PTU) is commonly used for inhibiting melanization of zebrafish embryos. In this study, the standard treatment with 0.2 mM PTU was demonstrated to specifically reduce eye size in larval fish starting at three days post-fertilization. This effect is likely the result of a reduction in retinal and lens size of PTU-treated eyes and is not related to melanization inhibition. This is because the eye size of tyr, a genetic mutant of tyrosinase whose activity is inhibited in PTU treatment, was not reduced. As PTU contains a thiocarbamide group which is presented in many goitrogens, suppressing thyroid hormone production is a possible mechanism by which PTU treatment may reduce eye size. Despite the fact that thyroxine level was found to be reduced in PTU-treated larvae, thyroid hormone supplements did not rescue the eye size reduction. Instead, treating embryos with six goitrogens, including inhibitors of thyroid peroxidase (TPO) and sodium-iodide symporter (NIS), suggested an alternative possibility. Specifically, three TPO inhibitors, including those that do not possess thiocarbamide, specifically reduced eye size; whereas none of the NIS inhibitors could elicit this effect. These observations indicate that TPO inhibition rather than a general suppression of thyroid hormone synthesis is likely the underlying cause of PTU-induced eye size reduction. Furthermore, the tissue-specific effect of PTU treatment might be mediated by an eye-specific TPO expression. Compared with treatment with other tyrosinase inhibitors or bleaching to remove melanization, PTU treatment remains the most effective approach. Thus, one should use caution when interpreting results that are obtained from PTU-treated embryos