Midkine-A functions upstream of Id2a to regulate cell cycle kinetics in the developing vertebrate retina

Abstract

Background: Midkine is a small heparin binding growth factor expressed in numerous tissues during development. The unique midkine gene in mammals has two paralogs in zebrafish: midkine-a (mdka) and midkine-b (mdkb). In the zebrafish retina, during both larval development and adult photoreceptor regeneration, mdka is expressed in retinal stem and progenitor cells and functions as a molecular component of the retina's stem cell niche. In this study, loss-of-function and conditional overexpression were used to investigate the function of Mdka in the retina of the embryonic zebrafish. Results: The results show that during early retinal development Mdka functions to regulate cell cycle kinetics. Following targeted knockdown of Mdka synthesis, retinal progenitors cycle more slowly, and this results in microphthalmia, a diminished rate of cell cycle exit and a temporal delay of cell cycle exit and neuronal differentiation. In contrast, Mdka overexpression results in acceleration of the cell cycle and retinal overgrowth. Mdka gain-of-function, however, does not temporally advance cell cycle exit. Experiments to identify a potential Mdka signaling pathway show that Mdka functions upstream of the HLH regulatory protein, Id2a. Gene expression analysis shows Mdka regulates id2a expression, and co-injection of Mdka morpholinos and id2a mRNA rescues the Mdka loss-of-function phenotype. Conclusions: These data show that in zebrafish, Mdka resides in a shared Id2a pathway to regulate cell cycle kinetics in retinal progenitors. This is the first study to demonstrate the function of Midkine during retinal development and adds Midkine to the list of growth factors that transcriptionally regulate Id proteins.

CERKL Knockdown Causes Retinal Degeneration in Zebrafish

“…Our RT-PCR semiquantitative analysis was supported by available zebrafish RNA-seq data (Fig. 2A) [34], [35], [36], [37], [38].…”

Müller glia: Stem cells for generation and regeneration of retinal neurons in teleost fish

“…For example, two zebrafish homologs of the heparin-binding growth factor Midkine, midkine a ( mdka ) and midkine b , may be important for regulating cell cycle dynamics during retinal regeneration. During retinal development, mdka positively regulates the transcriptional repressor id2a (inhibitor of DNA-binding 2a) , which modulates cell cycle progression from S- to M-phase without altering cell cycle timing or exit (Luo et al, 2012), and both midkines are re-expressed in proliferating Müller glia and photoreceptor progenitors after acute light lesion (Calinescu et al, 2009). The growth factor CNTF has dual roles in regeneration.…”

The midkine family of growth factors: diverse roles in nervous system formation and maintenance

“…Mdk and Ptn are proteins rich in basic aminoacids, with evolutionarily highly conserved cysteine residues. In zebrafish, the duplicated mdka and mdkb genes encode proteins that share 65% sequence identity (62.3% and 58.2% identity to human Mdk, respectively; mature protein without signal peptide), but show significant functional divergence during nervous system formation (Table ; Schafer et al ., ; Liedtke and Winkler, ) and in the developing retina (Luo et al ., ). This divergence offers a unique opportunity to analyse in vivo the effects of single amino acid substitutions in the two related sequences on protein structure and ultimately, biological function.…”