Importance of Intrinsic Mechanisms in Cell Fate Decisions in the Developing Rat Retina

MRC Laboratory for Molecular Cell Biology and Cell Biology Unit, University College London, London, WC1E 6BT United Kingdom.
Neuron (Impact Factor: 15.98). 01/2004; 40(5):897-904. DOI: 10.1016/S0896-6273(03)00756-6
Source: PubMed

ABSTRACT Cell diversification in the developing nervous system is thought to involve both cell-intrinsic mechanisms and extracellular signals, but their relative importance in particular cell fate decisions remains uncertain. In the mammalian retina, different cell types develop on a predictable schedule from multipotent retinal neuroepithelial cells (RNECs). A current view is that RNECs pass through a series of competence states, progressively changing their responsiveness to instructive extracellular cues, which also change over time. We show here, however, that embryonic day 16-17 (E16-17) rat RNECs develop similarly in serum-free clonal-density cultures and in serum-containing retinal explants--in the number of times they divide, the cell types they generate, and the order in which they generate these cell types. These surprising results suggest that extracellular signals may be less important than currently believed in determining when RNECs stop dividing and what cell types they generate when they withdraw from the cell cycle, at least from E16-17 onward.

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    • "Cell mixing experiments have shown that the differentiation potential of RPCs remains unchanged when they are placed in a temporally inappropriate environment (Watanabe and Raff, 1990; Altshuler and Cepko, 1992; Lillien and Cepko, 1992; Belliveau and Cepko, 1999). Moreover, when RPCs are plated at clonal density, they generate clones that are of similar size and composition as clones that develop in situ, and the general order of cell type production in the population is reproduced (Cayouette et al., 2003). Although feedback inhibition signals from the environment can clearly influence the proportion of specific cell types produced (Reh and Tully, 1986; Reh, 1992; Waid and McLoon, 1998; Wang et al., 2005), even in clonal cultures, these results are consistent with a model in which RPCs do not require specific instructive environmental signals for proper cell fate specification, but rather depend on cell intrinsic cues acting to bias their developmental potential over time (Cayouette et al., 2006; Gomes et al., 2011). "
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    ABSTRACT: BACKGROUND: We showed previously that the transcription factor Ikaros is expressed in early but not late retinal progenitors cells (RPCs), and is necessary and sufficient for the production of early-born neurons. Preliminary evidence using retinal explant cultures qualitatively suggested that Ikaros-positive RPCs might share a common lineage with Ikaros-negative RPCs. RESULTS: To explore further this question in vivo in a quantitative manner, we generated BAC transgenic mouse lines expressing Cre recombinase under the regulatory elements of the Ikaros gene, and crossed them with Cre reporter lines. Different transgenic lines labeled a variable number of RPCs, resulting in either dense or sparse radial arrays of reporter-positive progenies. Analysis of over 800 isolated cell arrays, which are most likely clones, confirmed that Ikaros-expressing RPCs generate both early- and late-born cell types in the same lineage, and that the overall cell composition of the arrays closely resembles that of the population of the mature retina. Interestingly, another sparse line did not label arrays, but appeared to specifically reflect Ikaros postmitotic expression in amacrine and ganglion cells. CONCLUSIONS: These mouse lines confirm the unbiased potential of the Ikaros lineage in vivo and provide novel tools for clonal lineage tracing and single neuron tracking in the retina.
    Developmental Dynamics 12/2012; 241(12). DOI:10.1002/dvdy.23881 · 2.67 Impact Factor
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    • "This data suggests the existence of a small population of progenitors passing slowly through G2/M. Live cell imaging indicates that M phase occurs relatively fast ($30 min) (Cayouette et al., 2003; Gomes et al., 2011), therefore the prolonged labeling of mitotic cells likely reflects variable G2 length. In prior cumulative labeling studies, the time needed to label approximately 90% of progenitors was equivalent to the time needed to label the remaining progenitors (Denham, 1967; Young, 1985b; Alexiades and Cepko, 1996), supporting the notion of a slow dividing RPC population. "
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    ABSTRACT: Background: Knowledge of gene expression kinetics around neuronal cell birth is required to dissect mechanisms underlying progenitor fate. Here, we timed cell cycle and neuronal protein silencing/induction during cell birth in the developing murine retina. Results: The pan-cell cycle markers Pcna and Mcm6 were present in the post-mitotic ganglion cell layer. Although confined to the neuroblastic layer (NBL), 6-7% of Ki67(+) cells lacked six progenitor/cell cycle markers, and expressed neuronal markers. To define protein extinction/induction timing, we defined G2/M length throughout retinogenesis, which was typically 1-2 h, but <10% cells took double this time. BrdU-chase analyses revealed that at E12.5, Tubb3 (Tuj1) appeared at M-phase, followed by Calb2 and Dcx at ∼2 h, Elavl2/3/4 at ∼4 h, and Map2 at ∼6 h after cell birth, and these times extended with embryonic age. Strikingly, Ki67 was not extinguished until up to a day after cell cycle exit, coinciding with exit from the NBL and induction of late markers such as Map1b/Uchl1/Rbfox3. Conclusions: A minor population of progenitors transits slowly through G2/M and, most importantly, some cell cycle proteins are retained for an unexpectedly long period in post-mitotic neurons. The high-resolution map of cell birth kinetics reported here provides a framework to better define mechanisms that regulate neurogenesis. Developmental Dynamics 241:1525-1544, 2012. © 2012 Wiley Periodicals, Inc.
    Developmental Dynamics 10/2012; 241(10):1525-44. DOI:10.1002/dvdy.23840 · 2.67 Impact Factor
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    • "How CNS structures, like the retina, of predictable sizes and cellular compositions arise from such variable lineages is a major unresolved question in developmental neuroscience. The variability of clones is an intrinsic cellular feature of RPCs (Cayouette et al., 2003). This is known because isolated rat RPCs grown in vitro produce clones of various sizes and compositions. "
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    ABSTRACT: A fundamental question in developmental neuroscience is how a collection of progenitor cells proliferates and differentiates to create a brain of the appropriate size and cellular composition. To address this issue, we devised lineage-tracing assays in developing zebrafish embryos to reconstruct entire retinal lineage progressions in vivo and thereby provide a complete quantitative map of the generation of a vertebrate CNS tissue from individual progenitors. These lineage data are consistent with a simple model in which the retina is derived from a set of equipotent retinal progenitor cells (RPCs) that are subject to stochastic factors controlling lineage progression. Clone formation in mutant embryos reveals that the transcription factor Ath5 acts as a molecular link between fate choice and mode of cell division, giving insight into the elusive molecular mechanisms of histogenesis, the conserved temporal order by which neurons of different types exit the cell cycle.
    Neuron 09/2012; 75(5):786-98. DOI:10.1016/j.neuron.2012.06.033 · 15.98 Impact Factor
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