LIM homeobox 9 (Lhx9) is a member of the LIM homeodomain transcription factor family, which expresses and functions in various vertebrate tissues, such as the gonads and pineal gland. Previous studies on lhx9 in zebrafish have mainly focused on the brain. However, little is known about the expression pattern of lhx9 during embryogenesis. Here, we detected lhx9 expression in zebrafish embryos using whole-mount in situ hybridization and found lhx9 expressed in heart, pectoral fin, and retina during their development in zebrafish. We then detailed the expression of lhx9 in retinal development. To further investigate the function of Lhx9 in retinogenesis, we performed morpholino (MO) knockdown experiments and found that upon lhx9 knockdown by MO, larvae presented normal eye development, retinal neural development, differentiation, proliferation, apoptosis, and responses to light stimulus. We not only elaborated the expression pattern of lhx9 in zebrafish embryogenesis, but we also demonstrated that lhx9 knockdown by morpholino does not affect the zebrafish retinal development, and our study provides data for further understanding of the role of Lhx9 in zebrafish retinal development.

LIM homeobox 4 (LHX4) is expressed in the photoreceptors (PRs) of the outer nuclear layer (ONL) and bipolar cells (BCs) of the inner nuclear layer (INL) in mouse and chicken retina. It regulates the subtype-specific development of rod BCs and cone BCs in the mouse retina. However, no report has been published on its expression and function in the zebrafish retina. In this study, we assessed the expression of Lhx4 using in situ hybridization (ISH) technique and explored its role in zebrafish (Danio rerio) retinal development via morpholino (MO) technology. We found that the expression of lhx4 in the zebrafish retina begins 48 h post-fertilization (hpf) and is continuously expressed in the ONL and INL. A zebrafish model constructed with lhx4 knockdown in the eyes through vivo-MO revealed that: lhx4 knockdown inhibits the differentiation of Parvalbumin+ amacrine cells (ACs) and Rhodopsin+ rod photoreceptors (RPs), enhances the expression of visual system homeobox 2 (vsx2); and damages the responses of zebrafish to light stimulus, without affecting the differentiation of OFF-BCs and rod BCs, and apoptosis in the retina. These findings reveal that lhx4 regulates neural differentiation in the retina and visual function during zebrafish embryonic development.

Nagashima-type palmoplantar keratosis (NPPK) is an autosomal recessive disorder with the main clinical manifestations of erythema, scales and keratotic plaques. In 2013, Kubo et al. 1 identified the causative gene as SERPINB7 by whole-exome sequencing (WES) in three unrelated Japanese NPPK patients. Currently, NPPK cases are mainly reported in Asian populations. Domestic and foreign studies have confirmed the founder effect of the c.796C>T mutation in NPPK. [2][3][4][5] The prevalence of NPPK based on large samples is estimated as 0.975/10 000 in the Chinese population. 6 SERPINB7 encodes the serine protease inhibitor subfamily B member 7 isoform, expressed explicitly in cells of the stratum corneum and stratum granulosum. 1,7 Although the causative gene of

Objective: The zinc finger, MIZ-type containing 1 (ZMIZ1) gene has been identified as a possible susceptibility gene associated with vitiligo, therefore we conducted this study to investigate the role of ZMIZ1 in pigmentation. Methods: We generate a zebrafish loss-of-function model using morpholino oligonucleotides (MOs), and two orthologs of human ZMIZ1 have been annotated (ZMIZ1a and ZMIZ1b). The expression profiles of ZMIZ1a and ZMIZ1b and their effects on the pigmentation in zebrafish were evaluated by using whole-mount in situ hybridization and melanin quantification. Statistical analysis was performed using the unpaired Student t-test or one-way analysis. Results: Investigation of the temporal and spatial expressions of these two transcripts suggested that the expressions of ZMIZ1a and ZMIZ1b in the brain start to emerge in a ubiquitous fashion from 2 days post-fertilization onwards. After the successful design and validation of MOs, we observed that ZMIZ1a and ZMIZ1b MOs caused embryonic developmental delays and malformations in zebrafish. Further analysis of the melanin content in the morphants revealed that ZMIZ1a significantly (49.1% for 0.667 mmol/L in ZMIZI1a group, P = 0.03) reduced the melanin content in a dose-dependent manner, but only the highest concentration of injected ZMIZ1b MOs significantly (50% for 0.667 mmol/L in ZMIZ1b group, P = 0.02) reduced the melanin content. A tyrosinase inhibition assay indicated no significant difference between the morphants and wild-type zebrafish. Conclusion: This study successfully modeled a susceptibility gene identified by genome-wide association studies in a zebrafish loss-of-function model and provides insights into the biological mechanism of pigmentation.

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