Transformation of cone precursors to functional rod photoreceptors by bZIP transcription factor NRL

Abstract

Networks of transcriptional regulatory proteins dictate specification of neural lineages from multipotent retinal progenitors. Rod photoreceptor differentiation requires the basic motifleucine zipper (bZIP) transcription factor NRL, because loss of Nrl in mice (Nrl ؊/؊ ) results in complete transformation of rods to functional cones. To examine the role of NRL in cell fate determination, we generated transgenic mice that express Nrl under the control of Crx promoter in postmitotic photoreceptor precursors of WT and Nrl ؊/؊ retina. We show that NRL expression, in both genetic backgrounds, leads to a functional retina with only rod photoreceptors. The absence of cones does not alter retinal lamination, although cone synaptic circuitry is now recruited by rods. Ectopic expression of NRL in developing cones can also induce rod-like characteristics and partially suppress cone-specific gene expression. We show that NRL is associated with specific promoter sequences in Thrb (encoding TR␤2 transcription factor required for M-cone differentiation) and S-opsin and may, therefore, directly participate in transcriptional suppression of cone development. Our studies establish that NRL is not only essential but is sufficient for rod differentiation and that postmitotic photoreceptor precursors are competent to make binary decisions during early retinogenesis.cell fate determination ͉ development ͉ gene regulation ͉ retina ͉ synaptic organization N euronal cell fate is determined by a hierarchical, stepwise process of binary decisions, commencing with multipotent progenitors that give rise to distinct cell lineages (1-3). The neural retina is an attractive model to investigate cell-fate determination; it contains seven major cell types that derive from common pool(s) of multipotent progenitor cells (4, 5). These retinal progenitors pass through sequential waves of competence, during which postmitotic cells can be specified to only a subset of neuronal fates (1, 6). Birth-dating studies in rodents indicate that ganglion cells, horizontal cells, cone photoreceptors, and amacrine cells are born prenatally, whereas most rod photoreceptors, bipolar cells, and Müller glia are generated postnatally (7-9). The orderly sequence of cell birth and a considerable overlap in their generation suggest a sequential program of cell intrinsic mechanisms and extrinsic signals that control cell fate decisions (10-16).Each neuronal lineage is meticulously established by highly coordinated transcription factor network(s) in response to local microenvironmental cues (17). Although extrinsic factors can promote differentiation (18,19), heterochronic mixing experiments demonstrate that progenitor cells at a particular time in development cannot be induced to generate temporally inappropriate cell types (1,20). Additionally, intrinsic priming of retinal progenitors appears to supersede the influence of environmental signals in specifying cell fate (21). Whether commitment of lineage-restricted precursors to a specific differentiation pathway is unidire...

Regulation of photoreceptor gene expression by Crx-associated transcription factor network

“…The inner neurons that ordinarily contact cones, such as ON cone bipolar and horizontal cells, make connections with rods in these mice. No cone-specific gene products were detected by RT-PCR or immunohistochemistry (Oh et al, 2007). Thus, Nrl is sufficient to guide photoreceptor precursors to a rod lineage.…”

“…However, conditional expression of Nrl later during S-cone differentiation, using the S-cone promoter on the Nrl −/− background, generated hybrid cells that co-expressed both rhodopsin and S-opsin. Although lineage-tracing experiments showed that some of the S-cones might have switched to a rod lineage, no rod function was detected by ERG in these mice (Oh et al, 2007), suggesting that differentiated photoreceptors may have limited plasticity, but require appropriate transcriptional regulation for maintenance. Another possible explanation, however, is that Nrl represses expression of S-cone genes, resulting in inactivation of cells already committed to the S-cone lineage.…”

“…To determine if Nrl is sufficient to induce the rod phenotype, Oh et al (2007) studied transgenic mice that express Nrl in all photoreceptor precursors under the control of a Crx promoter. This ectopic Nrl expression converts the retina of either wild-type (WT) or Nrl −/− mice to an allrod retina.…”

“…Chromatin immunoprecipitation assays showed that Nrl can directly bind to cone gene promoters, including S-opsin, M-opsin, cone arrestin (Arr3), and the cone transcription factor Trβ2 [ Oh et al, 2007) and Figure 3], suggesting that Nrl may directly regulate cone gene expression in rods. On the other hand, transient co-transfection assays with luciferase reporter constructs driven by either M-cone or S-cone opsin promoters showed that Nrl actually activates cone opsin promoters in vitro .…”

Retinoid-related orphan nuclear receptor RORβ is an early-acting factor in rod photoreceptor development

“…Nrl induces Nr2e3 expression and these two genes define a transcriptional hierarchy for rod differentiation. Nrl Ϫ/Ϫ mice overproduce S cones at the expense of rods (6) whereas ectopic Nrl expression converts cones to rods (14). Nr2e3 deficiency causes an enhanced cone phenotype and misexpression of cone genes in rods, suggesting that Nr2e3 represses cone genes to maintain the rod phenotype (15)(16)(17)(18)(19).…”