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Functional Diversification of the Four MARCKS Family Members in Zebrafish Neural Development
Daniel Prieto1, Flavio R Zolessi1,2
1Facultad de Ciencias, Sección Biología Celular, Universidad de la República, Montevideo, Uruguay.
Abstract:
Myristoylated alanin-rich C-kinase substrate (MARCKS) and MARCKS-like 1, each encoded by a different gene, comprise a very small family of actin-modulating proteins with essential roles in mammalian neural development. We show here that four genes (two marcks and two marcksl1) are present in teleosts including zebrafish, while ancient actinopterigians, sarcopterigian fishes, and chondrichtyans only have two. No marcks genes were found in agnaths or invertebrates. All four zebrafish genes are expressed during development, and we show here how their early knockdown causes defects in neural development, with some phenotypical differences. Knockdown of marcksa generated embryos with smaller brain and eyes, while marcksb caused different morphogenetic defects, such as larger hindbrain ventricle and folded retina. marcksl1a and marcksl1b morpholinos also caused smaller eyes and brain, although marcksl1a alone generated larger brain ventricles. At 24 hpf, marcksb caused a wider angle of the hindbrain walls, while marcksl1a showed a "T-shaped" neural tube and alterations in neuroepithelium organization. The double knockdown surprisingly produced new features, which included an increased neuroepithelial disorganization and partial neural tube duplications evident at 48 hpf, suggesting defects in convergent extension. This disorganization was also evident in the retina, although retinal ganglion cells were still able to differentiate. marcksl1b morphants presented a unique retinal phenotype characterized by the occurrence of sporadic ectopic neuronal differentiation. Although only marcksl1a morphant had a clear "ciliary phenotype," all presented significantly shorter cilia. Altogether, our data show that all marcks genes have functions in zebrafish neural development, with some differences that suggest the onset of protein diversification.
Insights
Myristoylated alanin-rich C-kinase substrate (MARCKS) and MARCKS-like 1 (MARCKSL1) genes are crucial for neural development. Zebrafish possess four MARCKS/MARCKSL1 genes, and their knockdown causes distinct neural and retinal defects, indicating functional diversification.
Area of Science:
- Developmental Biology
- Neuroscience
- Evolutionary Biology
Background:
- Myristoylated alanin-rich C-kinase substrate (MARCKS) and MARCKS-like 1 (MARCKSL1) are actin-modulating proteins vital for mammalian neural development.
- These proteins form a small gene family, with varying gene numbers across vertebrate evolution.
Purpose of the Study:
- To investigate the evolutionary presence and developmental roles of MARCKS and MARCKSL1 genes in teleosts, specifically zebrafish.
- To characterize the specific functions and potential diversification of the four zebrafish MARCKS/MARCKSL1 genes during neural development.
Main Methods:
- Comparative genomics to identify MARCKS and MARCKSL1 gene families across different vertebrate groups.
- Zebrafish morpholino-mediated knockdown of individual and combined marcks and marcksl1 genes.
- Phenotypic analysis of neural tube, brain, eye, retina, and cilia development at various embryonic stages.
Main Results:
- Zebrafish possess four MARCKS/MARCKSL1 genes (two marcks, two marcksl1), unlike fewer copies in older fish lineages or invertebrates.
- Individual gene knockdown resulted in specific neural defects, including altered brain size, ventricle morphology, retinal folding, and neuroepithelial disorganization.
- Double knockdown of marcksl1 genes led to enhanced neuroepithelial disorganization and neural tube duplications, suggesting roles in convergent extension.
- All morphants exhibited shorter cilia, with marcksl1a morphants showing a distinct ciliary phenotype.
Conclusions:
- All four zebrafish marcks and marcksl1 genes are expressed and play essential, albeit partially distinct, roles in neural development.
- The observed phenotypic differences suggest functional specialization and the early stages of protein diversification within this gene family in teleosts.
- These findings provide insights into the evolutionary conservation and divergence of actin-modulating proteins in vertebrate neural development.
