Dataset Information

Stathmin-like 4 is critical for the maintenance of neural progenitor cells in dorsal midbrain of zebrafish larvae.

ABSTRACT: A delicate balance between proliferating and differentiating signals is necessary to ensure proper growth and neuronal specification. By studying the developing zebrafish brain, we observed a specific and dynamic expression of a microtubule destabilizer gene, stathmin-like 4 (stmn4), in the dorsal midbrain region. The expression of stmn4 was mutually exclusive to a pan-neuronal marker, elavl3 that indicates its role in regulating neurogenesis. We showed the knockdown or overexpression of stmn4 resulted in premature neuronal differentiation in dorsal midbrain. We also generated stmn4 maternal-zygotic knockout zebrafish by the CRISPR/Cas9 system. Unexpectedly, only less than 10% of stmn4 mutants showed similar phenotypes observed in that of stmn4 morphants. It might be due to the complementation of the increased stmn1b expression observed in stmn4 mutants. In addition, time-lapse recordings revealed the changes in cellular proliferation and differentiation in stmn4 morphants. Stmn4 morphants displayed a longer G₂ phase that could be rescued by Cdc25a. Furthermore, the inhibition of Wnt could reduce stmn4 transcripts. These results suggest that the Wnt-mediated Stmn4 homeostasis is crucial for preventing dorsal midbrain from premature differentiation via the G₂ phase control during the neural keel stage.

SUBMITTER: Lin MJ

PROVIDER: S-EPMC5098158 | biostudies-literature | 2016 Nov

REPOSITORIES: biostudies-literature

ACCESS DATA

Publications

Stathmin-like 4 is critical for the maintenance of neural progenitor cells in dorsal midbrain of zebrafish larvae.

Lin Meng-Ju MJ Lee Shyh-Jye SJ

Scientific reports 20161107

A delicate balance between proliferating and differentiating signals is necessary to ensure proper growth and neuronal specification. By studying the developing zebrafish brain, we observed a specific and dynamic expression of a microtubule destabilizer gene, stathmin-like 4 (stmn4), in the dorsal midbrain region. The expression of stmn4 was mutually exclusive to a pan-neuronal marker, elavl3 that indicates its role in regulating neurogenesis. We showed the knockdown or overexpression of stmn4 r ...[more]

PMID: 27819330

Similar Datasets

Project description:Human pluripotent stem cells can be differentiated into midbrain dopaminergic (mDA) neurons by directing cells through a floor plate progenitor stage. The developmental identity of mDA neurons produced using floor plate protocols is similar to substantia nigra neurons, and this has improved the ability to model Parkinson's disease (PD) in a dish. Combined with the unlimited growth potential of pluripotent stem cells, mDA neural progenitor cell production can provide a scalable source of human dopaminergic (DA) neurons for diverse applications. However, due to the complexity and length of the protocols and inherent differences between cell lines, considerable variability of the final population of neurons is often observed. One solution to this problem is to cryopreserve committed mDA neural progenitor cells in a ready-to-use format. Creating a bank of cryopreserved mDA neural progenitor cells poised for neuronal differentiation could significantly improve reproducibility and facilitate collaborations. Here we have compared six (6) different commercial cryopreservation media and different freezing conditions for mDA neural progenitor cells differentiated from human embryonic stem cell (hESC) lines. Significant differences in cell recovery were observed at 24 h post-thawing, but no differences were observed immediately upon thawing. The presence of ROCK inhibitors improved cell recovery at 24 h for all cryopreservation media tested. A faster cooling rate of 1-2°C/min was significantly better than 0.5°C/min for all conditions tested, while rapid thawing at 37°C was not always superior to slow thawing at 4°C. Importantly, cryopreservation of mDA neural progenitor cells did not alter their potential to resume differentiation into mDA neurons. Banks of cryopreserved committed mDA neural progenitor cells provide a method to generate human DA neurons with reduced batch-to-batch variability, and establish a mechanism to share lineage-primed cells for collaborative research.

Project description:BackgroundThe Dmbx1 gene is important for the development of the midbrain and hindbrain, and mouse gene targeting experiments reveal that this gene is required for mediating postnatal and adult feeding behaviours. A single Dmbx1 gene exists in terrestrial vertebrate genomes, while teleost genomes have at least two paralogs. We compared the loss of function of the zebrafish dmbx1a and dmbx1b genes in order to gain insight into the molecular mechanism by which dmbx1 regulates neurogenesis, and to begin to understand why these duplicate genes have been retained in the zebrafish genome.ResultsUsing gene knockdown experiments we examined the function of the dmbx1 gene paralogs in zebrafish, dmbx1a and dmbx1b in regulating neurogenesis in the developing retina and midbrain. Dose-dependent loss of dmbx1a and dmbx1b function causes a significant reduction in growth of the midbrain and retina that is evident between 48-72 hpf. We show that this phenotype is not due to patterning defects or persistent cell death, but rather a deficit in progenitor cell cycle exit and differentiation. Analyses of the morphant retina or anterior hindbrain indicate that paralogous function is partially diverged since loss of dmbx1a is more severe than loss of dmbx1b. Molecular evolutionary analyses of the Dmbx1 genes suggest that while this gene family is conservative in its evolution, there was a dramatic change in selective constraint after the duplication event that gave rise to the dmbx1a and dmbx1b gene families in teleost fish, suggestive of positive selection. Interestingly, in contrast to zebrafish dmbx1a, over expression of the mouse Dmbx1 gene does not functionally compensate for the zebrafish dmbx1a knockdown phenotype, while over expression of the dmbx1b gene only partially compensates for the dmbx1a knockdown phenotype.ConclusionOur data suggest that both zebrafish dmbx1a and dmbx1b genes are retained in the fish genome due to their requirement during midbrain and retinal neurogenesis, although their function is partially diverged. At the cellular level, Dmbx1 regulates cell cycle exit and differentiation of progenitor cells. The unexpected observation of putative post-duplication positive selection of teleost Dmbx1 genes, especially dmbx1a, and the differences in functionality between the mouse and zebrafish genes suggests that the teleost Dmbx1 genes may have evolved a diverged function in the regulation of neurogenesis.

Dataset Information

Stathmin-like 4 is critical for the maintenance of neural progenitor cells in dorsal midbrain of zebrafish larvae.

Publications

Stathmin-like 4 is critical for the maintenance of neural progenitor cells in dorsal midbrain of zebrafish larvae.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets