C57BL/6J-Otud7a^em1Cpsc/J

Stock number: 031294
Product name: Otud7a knock-out (Otud7a null)
Research areas: Developmental Biology Research, Neurobiology Research

Description

Otud7a null mice recapitulate the main clinical neurodevelopmental aspects of human 15q13.3 microdeletion syndrome: developmental delay, vocal impairment, seizure-related abnormalities, motor deficits, hypotonia and acoustic startle deficit. When compared to the 15q13 deletion mouse model (all six 15q13.3 genes deleted), these Otud7a null mice exhibit the same cardinal phenotypes (with the exception of learning and memory deficits).

Detailed description

The proximal part of human chromosome 15 long (q) arm is susceptible to chromosomal deletions. Indeed, 15q13.3 microdeletions commonly encompass six protein-coding genes: FAN1, MTMR10, TRPM1, KLF13, OTUD7A and CHRNA7. 15q13.3 microdeletion syndrome is a rare condition (1 in 30,000-40,000) and is associated with the following neurodevelopmental defects (~incidence in affected individuals): intellectual disability (~55-75%), seizures/EEG abnormalities (~28-42%), language impairment/dysmorphia (~16-30%) and autism spectrum disorder (~10%) - as well as a wide spectrum of neuropsychiatric symptoms (ADHD, mood disorder, aggression, schizophrenia and/or hypotonia).

The Otud7a knock-out allele has most of the Otud7a OTU deubiquitinase domain deleted - creating a premature stop codon. Mice homozygous for this Otud7a knock-out allele (i.e., Otud7a null mice) recapitulate the main clinical neurodevelopmental aspects of human 15q13.3 microdeletion syndrome: developmental delay, vocal impairment, seizure-related abnormalities, motor deficits, hypotonia and acoustic startle deficit.
Specifically, Otud7a null mice exhibit a 30% weight reduction, a significant delay in developmental milestones (negative geotaxis, cliff aversion and incisor eruption/growth; but normal eye lid and ear opening), abnormal electrographic profile (repetitive-spike events and some seizure-like events associated with behavioral arrest), impaired ultrasonic vocalization, reduced acoustic startle response, various differences in social behaviors (may or may not correlate with autistic-like behaviors) and a significant impairment in motor coordination and motor learning (grip strength and rotarod).
Homozygous females have reduced prepulse inhibition (sensorimotor gating) - a feature impaired in schizophrenia patients.


Heterozygous mice are viable and fertile - also showing impaired ultrasonic vocalization and reduced acoustic startle response (both with less severity compared to homozygotes).


Breeding homozygotes has not been attempted to date (January 2018). Because homozygous pups are smaller in size, the donating investigator suggests placement of soft food pellets on the bottom of the cage to help pups thrive.

Otud7a null mice show no significant defects in overall brain structure/organization (including dendrite growth), memory and learning as adults (fear conditioning and novel object recognition) or open field activity, and they perform like wildtype mice on relevant assays that test for ADHD, anxiety or depression.

In summary, Otud7a null mice recapitulate the main clinical aspects of human 15q13.3 microdeletion syndrome. When compared to the 15q13 deletion mouse model having all six 15q13 genes deleted [Df(h15q13) ; PMID:27459725], these Otud7a null mice exhibit all the same cardinal phenotypes - with the exception of learning and memory deficits.

Development

The Otud7a knock-out allele was created in the laboratory of Dr. Christian P. Schaaf (Baylor College of Medicine) using CRISPR/Cas9 genome engineering in C57BL/6J mouse zygotes. Specifically, two single guide RNA (sgRNA) sequences were designed to delete a portion of exon 4, all of exon 5 and a portion of exon 6 of the OTU domain containing 7A locus (Otud7a) on chromosome 7. Deletion of that region (encoding most of the OTU deubiquitinase domain) creates a premature stop codon in Otud7a exon 8.
Founder mice with the Otud7a knock-out allele were identified by PCR (deletions at the desired locus and of the expected size). Founders were bred to C57BL/6J mice for germline transmission, and then at least one additional backcross to C57BL/6J was performed. No off-target mutations were found (via sequencing the top fifteen off-target candidates for each sgRNA predicted by CRISPR design tool). The Otud7a knock-out colony was then maintained by breeding heterozygous females with heterozygous males for several generations. In 2018, males with black coat color were sent to The Jackson Laboratory Repository. Upon arrival, sperm was cryopreserved. To establish our live colony, an aliquot of frozen sperm was used to fertilize C57BL/6J oocytes (Stock No. 000664).

Allele

Symbol: Otud7a^em1Cpsc
Name: endonuclease-mediated mutation 1, Christian Schaaf
MGI accession ID: MGI:6108137
Generation method: Endonuclease-mediated
Attributes: Null/Knockout
Marker symbol: Otud7a
Marker name: OTU domain containing 7A
Marker synonyms: C15orf16; C16ORF15; Cezanne 2 protein; CEZANNE2; NEDHS; Otud7; RGD1563721
Chromosome: 7
Strain of origin: C57BL/6J
Molecular note: Two single guide RNA (sgRNA) sequences were designed to delete a portion of exon 4, all of exon 5 and a portion of exon 6 of the gene. Deletion of that region (encoding most of the OTU deubiquitinase domain) creates a premature stop codon in exon 8. No off-target mutations were found (via sequencing the top fifteen off-target candidates for each sgRNA predicted by CRISPR design tool).