Abstract:Background
The cell cycle regulator Cyclin D1 is expressed in embryonic retinal progenitor cells (RPCs) and regulates their cell cycle rate and neurogenic output. We report here that Cyclin D1 also has important functions in postnatal retinal histogenesis.
Results
The initial production of Müller glia and bipolar cells was enhanced in Cyclin D1 knockout (Ccnd1−/−) retinas. Despite a steeper than normal rate of depletion of the RPC population at embryonic ages, postnatal Ccnd1−/− retinas exhibited an extended… Show more
“…Fig. S3D) and adult mouse retinas (Das et al, ; Osakada et al, ). Interestingly, in EGF‐stimulated retinas Ccnd3 remains expressed and even more Ccnd3+ are generated until DEV12 (Supp.…”
The mechanisms limiting neuronal regeneration in mammals and their relationship with reactive gliosis are unknown. Müller glia (MG), common to all vertebrate retinas, readily regenerate neuron loss in some species, but normally not in mammals. However, experimental stimulation of limited mammalian retina regeneration has been reported. Here, we use a mouse retina organ culture approach to investigate the MG responses at different mouse ages. We found that MG undergo defined spatio-temporal changes upon stimulation. In EGF-stimulated juvenile postmitotic retinas, most MG upregulate cell-cycle regulators (Mcm6, Pcna, Ki67, Ccnd1) within 48 h ex vivo; some also express the neurogenic factors Ascl1, Pax6, and Vsx2; up to 60% re-enter the cell cycle, some of which delaminate to divide mostly apically; and the majority cease to proliferate after stimulation. A subpopulation of MG progeny starts to express transcription factors (Ptf1a, Nr4a2) and neuronal (Calb1, Calb2, Rbfox3), but not glial, markers, indicating neurogenesis. BrdU-tracking, genetic lineage-tracing, and transgenic-reporter experiments suggest that MG reprogram to a neurogenic stage and proliferate; and that some MG progeny differentiate into neuronal-like cells, most likely amacrines, no photoreceptors; most others remain in a de-differentiated state. The mouse MG regeneration potential becomes restricted, dependent on the age of the animal, as observed by limited activation of the cell cycle and neurogenic factors. The stage-dependent analysis of mouse MG revealed similarities and differences when compared with MG-derived regeneration in fish and chicks. Therefore, the mouse retina ex vivo approach is a potential assay for understanding and overcoming the limitations of mammalian MG-derived neuronal regeneration. Postmitotic MG in mouse retina ex vivo can be stimulated to proliferate, express neurogenic factors, and generate progeny expressing neuronal or glial markers. This potential regenerative competence becomes limited with increasing mouse age.
“…Fig. S3D) and adult mouse retinas (Das et al, ; Osakada et al, ). Interestingly, in EGF‐stimulated retinas Ccnd3 remains expressed and even more Ccnd3+ are generated until DEV12 (Supp.…”
The mechanisms limiting neuronal regeneration in mammals and their relationship with reactive gliosis are unknown. Müller glia (MG), common to all vertebrate retinas, readily regenerate neuron loss in some species, but normally not in mammals. However, experimental stimulation of limited mammalian retina regeneration has been reported. Here, we use a mouse retina organ culture approach to investigate the MG responses at different mouse ages. We found that MG undergo defined spatio-temporal changes upon stimulation. In EGF-stimulated juvenile postmitotic retinas, most MG upregulate cell-cycle regulators (Mcm6, Pcna, Ki67, Ccnd1) within 48 h ex vivo; some also express the neurogenic factors Ascl1, Pax6, and Vsx2; up to 60% re-enter the cell cycle, some of which delaminate to divide mostly apically; and the majority cease to proliferate after stimulation. A subpopulation of MG progeny starts to express transcription factors (Ptf1a, Nr4a2) and neuronal (Calb1, Calb2, Rbfox3), but not glial, markers, indicating neurogenesis. BrdU-tracking, genetic lineage-tracing, and transgenic-reporter experiments suggest that MG reprogram to a neurogenic stage and proliferate; and that some MG progeny differentiate into neuronal-like cells, most likely amacrines, no photoreceptors; most others remain in a de-differentiated state. The mouse MG regeneration potential becomes restricted, dependent on the age of the animal, as observed by limited activation of the cell cycle and neurogenic factors. The stage-dependent analysis of mouse MG revealed similarities and differences when compared with MG-derived regeneration in fish and chicks. Therefore, the mouse retina ex vivo approach is a potential assay for understanding and overcoming the limitations of mammalian MG-derived neuronal regeneration. Postmitotic MG in mouse retina ex vivo can be stimulated to proliferate, express neurogenic factors, and generate progeny expressing neuronal or glial markers. This potential regenerative competence becomes limited with increasing mouse age.
“…We next asked how RPC proliferation is supported in the absence of SOX2. D-type Cyclins are expressed in RPCs and promote progression through the cell cycle [ 51 – 53 ]. We examined expression of CyclinD1 and CyclinD3 protein by immunofluorescence in control and mutant OCs at E16.5 and P0.…”
Section: Resultsmentioning
confidence: 99%
“…CyclinD3 protein was confined to the prospective CE of controls but expanded into the central OC of mutants (Figure 7 G, H versus J, K). Co-labeling with CyclinD1 and CyclinD3 indicated that these two proteins, which are normally mutually exclusive, were aberrantly co-expressed in some Sox2 -depleted cells (Figure 7 M-P; arrows) [ 53 ]. Figure 7 D- type Cyclins are aberrantly expressed in Sox2- ablated optic cup progenitor cells (OCPCs) at E16.5.…”
BackgroundEye development in vertebrates relies on the critical regulation of SOX2 expression. Humans with mutations in SOX2 often suffer from eye defects including anophthalmia (no eye) and microphthalmia (small eye). In mice, deletion of Sox2 in optic cup progenitor cells results in loss of neural competence and cell fate conversion of the neural retina to a non-neurogenic fate, specifically the acquisition of fate associated with progenitors of the ciliary epithelium. This fate is also promoted with constitutive expression of stabilized β-Catenin in the optic cup, where the WNT pathway is up-regulated. We addressed whether SOX2 co-ordinates the neurogenic boundary of the retina through modulating the WNT/β-Catenin pathway by using a genetic approach in the mouse.ResultsUpon deletion of Sox2 in the optic cup, response to WNT signaling was expanded, correlating with loss of neural competence, cell fate conversion of the neural retina to ciliary epithelium primordium and, in addition, increased cell cycle time of optic cup progenitors. Removal of Ctnnb1 rescued the cell fate conversion; however, the loss of neural competence and the proliferation defect resulting from lack of SOX2 were not overcome. Lastly, central Sox2-deficient optic cup progenitor cells exhibited WNT-independent up-regulation of D-type Cyclins.ConclusionWe propose two distinct roles for SOX2 in the developing retina. Our findings suggest that SOX2 antagonizes the WNT pathway to maintain a neurogenic fate and, in contrast, regulates cycling of optic cup progenitors in a WNT-independent manner. Given that WNT signaling acting upstream of SOX2 has been implicated in the tumorigenicity of embryonic stem cell-derived retinal progenitor cells, our results distinguish the endogenous role of WNT signaling in early optic cup patterning and support a WNT-independent role for SOX2 in maintaining retinal progenitor cell proliferation.Electronic supplementary materialThe online version of this article (doi:10.1186/1749-8104-9-27) contains supplementary material, which is available to authorized users.
“…Consistent with this explanation are the phenotypes reported in several cell cycle mutants that alter the dynamics of cell cycle progression. For example, loss of cyclin D1, a protein that controls cell cycle transition from G1 to the S-phase, results in a loss of RGCs and an increase in the fate of late-born cells (12). Similarly, loss of cyclin D2 alters the cell cycle duration of the cells within the ciliary marginal zone, resulting in a reduction in the ipsilateral and contralateral RGC population (32).…”
Section: Cell Cycle and Neurogenesismentioning
confidence: 99%
“…Thus, cell cycle components are able to drive the cellular processes during retinal neurogenesis and are key regulators of cell fate specification (9,10). Factors that promote cell cycle progression like cyclins and cyclin-dependent kinases are highly expressed in the RPCs, but their expression is down-regulated in the differentiated retinal cells, consistent with a function in maintaining cell proliferation (11)(12)(13). Conversely, cyclin-dependent kinase inhibitors are involved in promoting cell cycle exit, and their loss can result in increased proliferation (14,15).…”
A single pool of multipotent retinal progenitor cells give rise to the diverse cell types within the mammalian retina. Such cellular diversity is due to precise control of various cellular processes like cell specification, proliferation, differentiation, and maturation. Circadian clock genes can control the expression of key regulators of cell cycle progression and therefore can synchronize the cell cycle state of a heterogeneous population of cells. Here we show that the protein encoded by the circadian clock gene brain and muscle arnt‐like protein‐1 (Bmal1) is expressed in the embryonic retina and is required to regulate the timing of cell cycle exit. Accordingly, loss of Bmal1 during retinal neurogenesis results in increased S‐phase entry and delayed cell cycle exit. Disruption in cell cycle kinetics affects the timely generation of the appropriate neuronal population thus leading to an overall decrease in the number of retinal ganglion cells, amacrine cells, and an increase in the number of the late‐born type II cone bipolar cells as well as the Müller glia. Additionally, the mislocalized Müller cells are observed in the photoreceptor layer in the Bmal1 conditional mutants. These changes affect the functional integrity of the visual circuitry as we report a significant delay in visual evoked potential implicit time in the retina‐specific Bmal1 null animals. Our results demonstrate that Bmal1 is required to maintain the balance between the neural and glial cells in the embryonic retina by coordinating the timing of cell cycle entry and exit. Thus, Bmal1 plays an essential role during retinal neurogenesis affecting both development and function of the mature retina.—Sawant, O. B., Jidigam, V. K., Fuller, R. D., Zucaro, O. F., Kpegba, C., Yu, M., Peachey, N. S., Rao, S. The circadian clock gene Bmal1 is required to control the timing of retinal neurogenesis and lamination of Müller glia in the mouse retina. FASEB J. 33, 8745–8758 (2019). http://www.fasebj.org
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