Characterization of NHLRC2 gene-edited mice: a model for bovine developmental duplications

Developmental Duplications

Delhotal, Jocelyn Denae ; Dilger, Anna C., ; ,Wheeler, Matthew B. ; Beever ; Jonathan E.

关键词: Developmental Duplications;   

Others  :  https://www.ideals.illinois.edu/bitstream/handle/2142/92824/DELHOTAL-THESIS-2016.pdf?sequence=1&isAllowed=y

美国|英语

来源: The Illinois Digital Environment for Access to Learning and Scholarship

【 摘 要 】

Developmental duplications (DD) is a genetic condition recently characterized in Angus cattle. It is a congenital abnormality where duplication of neural crest derived tissues occurs during embryonic development. A common phenotypic presentation of the condition includes calves born with polymelia most frequently involving duplication of the front limbs that protrude from the neck or shoulder region. Aside from polymelia, DD affected individuals present malformations associated with neural tube defects (NTDs). Genome-wide association studies have identified a single locus associated with this disease phenotype. Further investigation has identified the putative mutation as a nonsynonymous substitution (p.Val311Ala) in the NHL repeat-containing 2 (NHLRC2) gene. Transcription activator-like effector nucleases (TALENs) targeting exon 5 of NHLRC2 were used for gene-editing of the orthologous locus in mice to further investigate the role of NHLRC2 in development. Three mouse lines were generated with mutations having varying impacts on the NHLRC2 protein. Two mutations, -2 bp and -19 bp, are predicted to cause a prematurely truncated protein and one mutation, -12 bp, the deletion of four amino acids, residues 307 through 310, adjacent to the corresponding bovine substitution. Heterozygous mice of each line were intermated to phenotypically characterize homozygous progeny. Genotyping of the offspring revealed absence of homozygous individuals suggesting embryonic lethality. Because initiation of neural tube closure in mice occurs at day E8.5, embryonic death was assessed at this developmental day by harvesting embryos from heterozygous matings at day E8.5 with subsequent genotyping. Again, no homozygous embryos could be detected, however yolk sacs containing no embryos were observed. Furthermore, the number of yolk sacs exceeded the average number of live births by 68% (11.3 vs. 6.7), indicating embryonic lethality of homozygous individuals most likely occurs between fertilization and E8.5. Thus, we suggest that NHLRC2 is essential during mammalian development and hypothesize NHLRC2 plays a significant role in neurulation.