Notch pathway mutants do not equivalently perturb mouse embryonic retinal development.

In the vertebrate eye, Notch ligands, receptors, and ternary complex components determine the destiny of retinal progenitor cells in part by regulating Hes effector gene activity. There are multiple paralogues for nearly every node in this pathway, which results in numerous instances of redundancy and compensation during development. To dissect such complexity at the earliest stages of eye development, we used seven germline or conditional mutant mice and two spatiotemporally distinct Cre drivers. We perturbed the Notch ternary complex and multiple Hes genes to understand if Notch regulates optic stalk/nerve head development; and to test intracellular pathway components for their Notch-dependent versus -independent roles during retinal ganglion cell and cone photoreceptor competence and fate acquisition. We confirmed that disrupting Notch signaling universally blocks progenitor cell growth, but delineated specific pathway components that can act independently, such as sustained Hes1 expression in the optic stalk/nerve head. In retinal progenitor cells, we found that among the genes tested, they do not uniformly suppress retinal ganglion cell or cone differentiation; which is not due differences in developmental timing. We discovered that shifts in the earliest cell fates correlate with expression changes for the early photoreceptor factor Otx2, but not with Atoh7, a factor required for retinal ganglion cell formation. During photoreceptor genesis we also better defined multiple and simultaneous activities for Rbpj and Hes1 and identify redundant activities that occur downstream of Notch. Given its unique roles at the retina-optic stalk boundary and cone photoreceptor genesis, our data suggest Hes1 as a hub where Notch-dependent and -independent inputs converge.

Not all Notch pathway mutations are equal in the embryonic mouse retina.

In the vertebrate retina, combinations of Notch ligands, receptors, and ternary complex components determine the destiny of retinal progenitor cells by regulating Hes effector gene activity. Owing to reiterated Notch signaling in numerous tissues throughout development, there are multiple vertebrate paralogues for nearly every node in this pathway. These Notch signaling components can act redundantly or in a compensatory fashion during development. To dissect the complexity of this pathway during retinal development, we used seven germline or conditional mutant mice and two spatiotemporally distinct Cre drivers. We perturbed the Notch ternary complex and multiple Hes genes with two overt goals in mind. First, we wished to determine if Notch signaling is required in the optic stalk/nerve head for Hes1 sustained expression and activity. Second, we aimed to test if Hes1, 3 and 5 genes are functionally redundant during early retinal histogenesis. With our allelic series, we found that disrupting Notch signaling consistently blocked mitotic growth and overproduced ganglion cells, but we also identified two significant branchpoints for this pathway. In the optic stalk/nerve head, sustained Hes1 is regulated independent of Notch signaling, whereas during photoreceptor genesis both Notch-dependent and -independent roles for Rbpj and Hes1 impact photoreceptor genesis in opposing manners.

son is necessary for proper vertebrate blood development.

The gene SON is on human chromosome 21 (21q22.11) and is thought to be associated with hematopoietic disorders that accompany Down syndrome. Additionally, SON is an RNA splicing factor that plays a role in the transcription of leukemia-associated genes. Previously, we showed that mutations in SON cause malformations in human and zebrafish spines and brains during early embryonic development. To examine the role of SON in normal hematopoiesis, we reduced expression of the zebrafish homolog of SON in zebrafish at the single-cell developmental stage with specific morpholinos. In addition to the brain and spinal malformations we also observed abnormal blood cell levels upon son knockdown. We then investigated how blood production was altered when levels of son were reduced. Decreased levels of son resulted in lower amounts of red blood cells when visualized with lcr:GFP transgenic fish. There were also reduced thrombocytes seen with cd41:GFP fish, and myeloid cells when mpx:GFP fish were examined. We also observed a significant decrease in the quantity of T cells, visualized with lck:GFP fish. However, when we examined their hematopoietic stem and progenitor cells (HSPCs), we saw no difference in colony-forming capability. These studies indicate that son is essential for the proper differentiation of the innate and adaptive immune system, and further investigation determining the molecular pathways involved during blood development should elucidate important information about vertebrate HSPC generation, proliferation, and differentiation.