N-myc regulates growth and fiber cell differentiation in lens development

Abstract

Myc proto-oncogenes regulate diverse cellular processes during development, but their roles during morphogenesis of specific tissues are not fully understood. We found that c-myc regulates cell proliferation in mouse lens development and previous genome-wide studies suggested functional roles for N-myc in developing lens. Here, we examined the role of N-myc in mouse lens development. Genetic inactivation of N-myc in the surface ectoderm or lens vesicle impaired eye and lens growth, while "late" inactivation in lens fibers had no effect. Unexpectedly, defective growth of N-myc--deficient lenses was not associated with alterations in lens progenitor cell proliferation or survival. Notably, N-myc-deficient lens exhibited a delay in degradation of DNA in terminally differentiating lens fiber cells. RNA-sequencing analysis of N-myc--deficient lenses identified a cohort of down-regulated genes associated with fiber cell differentiation that included DNaseIIβ. Further, an integrated analysis of differentially expressed genes in N-myc-deficient lens using normal lens expression patterns of iSyTE, N-myc-binding motif analysis and molecular interaction data from the String database led to the derivation of an N-myc-based gene regulatory network in the lens. Finally, analysis of N-myc and c-myc double-deficient lens demonstrated that these Myc genes cooperate to drive lens growth prior to lens vesicle stage. Together, these findings provide evidence for exclusive and cooperative functions of Myc transcription factors in mouse lens development and identify novel mechanisms by which N-myc regulates cell differentiation during eye morphogenesis.

The RNA-binding protein Celf1 post-transcriptionally regulates p27Kip1 and Dnase2b to control fiber cell nuclear degradation in lens development

“…Further, analysis of the Celf1 cKO/lacZKI lens DEGs by comparing them to normal mouse lens gene expression data in iSyTE shows that majority of the genes down-regulated in Celf1 cKO/lacZKI lenses exhibit highly enriched expression in lens development, while up-regulated genes had no such pattern ( Fig 2 B )–indicating that Celf1 is necessary for expression of genes associated with differentiating fiber cells. To identify high-priority targets among Celf1 cKO/lacZKI lens DEGs, we next applied an effective filtering criteria that in the past has successfully pointed to key genes that explained the lens phenotype in other gene knockout mice [ 18 , 22 – 24 ]. These analyses identify known–as well as new–candidate genes involved in different aspects of fiber cell differentiation, in turn offering avenues for detailed investigation (see below) for explaining the cataract pathology observed in Celf1 deficient lenses.…”

Transcriptome analysis of developing lens reveals abundance of novel transcripts and extensive splicing alterations

“…In contrast, expression profiles from E18 and post-natal stages exhibited lower correlation with respect to E15.5 dataset, further confirming the quality and robustness of expression profiles to delineate the developmental stages. Although, increasing number of studies using RNA-sequencing protocols are able to generate a wild type control as part of their research projects leading to stage-specific developmental transcriptomes 56 – 58 , several issues need to be considered before employing them in large-scale meta-analysis studies, which can significantly improve the quality and number of high confidence predictions. For instance, several of these publicly available datasets are generated with single replicates, provide separate transcriptomes of epithelial and fiber cells as opposed to whole lens, are generated with differing read lengths and arise from different labs.…”

De novogenesis of retinal ganglion cells by targeted expression of KLF4in vivo

“…In loss-of-function experiments, sections near of the optic nerve were. In gain-of-function experiments, regions with electroporated GFP+ cells were immunoreacted as reported previously (Cavalheiro et al, 2017). All images were acquired using structural illumination (Imager M2 Apotome Zeiss).…”