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Rice, D. S.

Publications and source records attributed to Rice, D. S..

2 recordsLinked to original sources

A spontaneous mutation in ADIPOR1 causes retinal degeneration in mice

Adiponectin receptor 1 (ADIPOR1) is a transmembrane protein necessary for normal anatomy and physiology in the retina. In a recent study of complement factor H knockout mice (Cfh-/-), our lab discovered a flecked retina phenotype and retinal thinning by fundus imaging and optical coherence tomography (OCT), respectively. The phenotype was observed in a subset (50%) of Cfh-/- mice. The thinning observed in vivo is due to an early degeneration of rod photoreceptors. This phenotype has not been reported in published studies of Cfh-/- mice. AdipoR1 knockout mice (AdipoR1-/-) and mice deficient in Membrane Frizzled Related Protein (MFRP) exhibit this phenotype, suggesting an involvement in the emergence of the retinal degeneration observed in a subset of Cfh-/- mice. Cfh and AdipoR1 are located in close proximity on mouse Chromosome 1 (Chr1) and a complementation cross between Cfh and AdipoR1 mice with retinal degeneration produced 100% progeny with retinal degeneration. Sequencing of the Cfh-/- mice revealed a c.841 C > T mutation in AdipoR1. Furthermore, one Cfh wildtype (of Cfh+/+) and 2 heterozygous (of Cfh+/-) mice exhibited retinal degeneration and were homozygous for the point mutation. The c.841 C > T mutation results in a proline to serine conversion at position 281 (P281S) in ADIPOR1. This residue is critical for ADIPOR1 open and closed conformations in the membrane. In silico modeling of candidate ADIPOR1 ligands, 11-cis-retinaldehyde and docosahexaenoic acid (DHA), that are deficient in AdipoR1-/-, suggests that ADIPOR1 is involved in trafficking retinoids and fatty acids and their combined deficiency in the ADIPOR1 mutant retinas might explain the retinal degeneration phenotype.

neuroscience↗

Highly efficient scarless knock-in of reporter genes into human and mouse pluripotent stem cells via transient antibiotic selection

Pluripotent stem cells (PSCs) edited with genetic reporters are useful tools for differentiation analysis and for isolation of specific cell populations for study. Reporter integration into the genome is now commonly achieved by targeted DNA nuclease-enhanced homology directed repair (HDR). However, human PSCs are known to have a low frequency of gene knock-in (KI) by HDR, making reporter line generation an arduous process. Here, we report a methodology for scarless KI of large fluorescent reporter genes into PSCs by transient selection with puromycin or zeocin. With this method, we can perform targeted KI of a single reporter gene with up to 65% efficiency, as well as simultaneous KI of two reporter genes into different loci with up to 11% efficiency. Additionally, we demonstrate that this method also works in mouse PSCs.

cell biology↗