C57BL/6NCrl-Maddem1(R327stop)Ccpcz
| Gene Symbol | Madd |
| Gene Name | MAP kinase activating death domain |
| Gene Synonym(s) | DENN, IG20, KIAA0358, RAB3GEP |
| ENSEMBL | human: ENSG00000110514 mouse: ENSMUSG00000040687 |
| Allele Symbol | C57BL/6NCrl-Maddem1(R327stop)Ccpcz |
| Allele Name | endonuclease-mediated allele 1, Ccpcz |
| Allele Type | point mutation, R327* (R327 stop) |
| Genomic location | human: Chromosome 11: 47,269,161-47,330,038 forward strand, GRCh38:CM000673.2 mouse: Chromosome 2: 90,967,705-91,014,182 reverse strand, GRCm39:CM000995.3 |
| Strain of Origin | C57Bl/6NCrl |
| Molecular Note | Mutation introduced by CRISPR/Cas9 double strand break and electroporation of dsDNA template with >40 bp homologous arms. Genotyped by PCR amplification of the sequence surrounding the mutation site and subsequent restriction digest by XhoI as well as Sanger sequencing. |
| Mutations Made by | Czech Centre of Phenogenomics, Institute of Molecular Genetics of the Czech Academy of Sciences |
Description
The MADD gene encodes a MAPK-activating protein that belongs to the DENN protein family, which regulates the Rab family is small GTPases. MADD acts as a guanine nucleotide exchange factor (GEF) for Rab3 (see 602536), which itself is present on synaptic vesicles and regulates neurotransmitter release. MADD also interacts with TNFR1 (191190), which stimulates intracellular signaling pathways and activation of transcription factors under stress conditions. The MADD gene undergoes extensive splicing and generates at least 7 different isoforms with variable tissue expression (summary by Schneeberger et al., 2020).
Associated pathology
DEVELOPMENTAL DELAY WITH ENDOCRINE, EXOCRINE, AUTONOMIC, AND HEMATOLOGIC ABNORMALITIES (DEEAH)
DEEAH syndrome is an autosomal recessive multisystemic disorder with onset in early infancy. Affected individuals usually present in the perinatal period with respiratory insufficiency, apneic episodes, and generalized hypotonia. The patients have failure to thrive and severely impaired global development with poor acquisition of motor, cognitive, and language skills. Other common features include endocrine, pancreatic exocrine, and autonomic dysfunction, as well as hematologic disturbances, mainly low hemoglobin. Patients also have dysmorphic and myopathic facial features. Additional more variable features include seizures, undescended testes, and distal skeletal anomalies. Death in early childhood may occur (summary by Schneeberger et al., 2020).
Model Development
We used synthetic ssODN carrying the target R327* mutation (CGA>TGA) plus one extra silent mutation (GTA>GTG) to destroy the PAM site. ssODN had homologous regions on both sides of the mutation >40 bp long. Used ssODN was premixed with Cas9 protein and gRNA and electroporated into zygotes. Zygotes or 2-cell stage embryos were transferred into pseudopregnant recipient mice. Resulting mice (G0 generation) were genotyped by PCR and subsequent restriction digest. The presence of R327* mutation and one extra silent mutation, and no other mutations in the vicinity of targeted region, were confirmed by Sanger sequencing. Positive individuals were backcrossed with C57BL/6NCrl for two generations to establish the colony.