Inhibitory G proteins play multiple roles to polarize sensory hair cell morphogenesis.

Inhibitory G alpha (GNAI or Gαi) proteins are critical for the polarized morphogenesis of sensory hair cells and for hearing. The extent and nature of their actual contributions remains unclear, however, as previous studies did not investigate all GNAI proteins and included non-physiological approaches. Pertussis toxin can downregulate functionally redundant GNAI1, GNAI2, GNAI3, and GNAO proteins, but may also induce unrelated defects.

Dystonin modifiers of junctional epidermolysis bullosa and models of epidermolysis bullosa simplex without dystonia musculorum.

The Lamc2jeb junctional epidermolysis bullosa (EB) mouse model has been used to demonstrate that significant genetic modification of EB symptoms is possible, identifying as modifiers Col17a1 and six other quantitative trait loci, several with strong candidate genes including dystonin (Dst/Bpag1). Here, CRISPR/Cas9 was used to alter exon 23 in mouse skin specific isoform Dst-e (Ensembl GRCm38 transcript name Dst-213, transcript ID ENSMUST00000183302.5, protein size 2639AA) and validate a proposed arginine/glutamine difference at amino acid p1226 in B6 versus 129 mice as a modifier of EB.

Functional analysis of Collagen 17a1: A genetic modifier of junctional epidermolysis bullosa in mice.

Previous work strongly implicated Collagen 17a1 (Col17a1) as a potent genetic modifier of junctional epidermolysis bullosa (JEB) caused by a hypomorphic mutation (Lamc2jeb) in mice. The importance of the noncollagenous domain (NC4) of COLXVII was suggested by use of a congenic reduction approach that restricted the modifier effect to 2-3 neighboring amino acid changes in that domain. The current study utilizes TALEN and CRISPR/Cas9 induced amino acid replacements and in-frame indels nested to NC4 to further investigate the role of this and adjoining COLXVII domains both as modifiers and primary risk effectors.

A novel HSPB1 mouse model of Charcot-Marie-Tooth Disease.

Charcot-Marie-Tooth Disease (CMT) is a commonly inherited peripheral polyneuropathy. Clinical manifestations for this disease include symmetrical distal polyneuropathy, altered deep tendon reflexes, distal sensory loss, foot deformities, and gait abnormalities. Genetic mutations in heat shock proteins have been linked to CMT2.

Genetically diverse mouse platform to xenograft cancer cells.

The lack of genetically diverse preclinical animal models in basic biology and efficacy testing has been cited as a potential cause of failure in clinical trials. We developed and characterized five diverse RAG1 null mouse strains as models that allow xenografts to grow. In these strains, we characterized the growth of breast cancer, leukemia and glioma cell lines.

Programmable RNA-Guided Large DNA Transgenesis by CRISPR/Cas9 and Site-Specific Integrase Bxb1.

The inability to insert large DNA constructs into the genome efficiently and precisely is a key challenge in genomic engineering. Random transgenesis, which is widely used, lacks precision, and comes with a slew of drawbacks. Lentiviral and adeno-associated viral methods are plagued by, respectively, DNA toxicity and a payload capacity of less than 5 kb.

Efficient targeted transgenesis of large donor DNA into multiple mouse genetic backgrounds using bacteriophage Bxb1 integrase.

The development of mouse models of human disease and synthetic biology research by targeted transgenesis of large DNA constructs represent a significant genetic engineering hurdle. We developed an efficient, precise, single-copy integration of large transgenes directly into zygotes using multiple mouse genetic backgrounds. We used in vivo Bxb1 mediated recombinase-mediated cassette exchange (RMCE) with a transgene "landing pad" composed of dual heterologous Bxb1 attachment (att) sites in cis, within the Gt(ROSA)26Sor safe harbor locus.

Inactive rhomboid proteins RHBDF1 and RHBDF2 (iRhoms): a decade of research in murine models.

Rhomboid proteases, first discovered in Drosophila, are intramembrane serine proteases. Members of the rhomboid protein family that are catalytically deficient are known as inactive rhomboids (iRhoms). iRhoms have been implicated in wound healing, cancer, and neurological disorders such as Alzheimer's and Parkinson's diseases, inflammation, and skin diseases.

Functional humanization of immunoglobulin heavy constant gamma 1 Fc domain human transgenic mice.

A major asset of many monoclonal antibody (mAb)-based biologics is their persistence in circulation. The MHC class I family Fc receptor, FCGRT, is primarily responsible for this extended pharmacokinetic behavior. Engagement of FCGRT with the crystallizable fragment (Fc) domain protects IgG from catabolic elimination, thereby extending the persistence and bioavailability of IgG and related Fc-based biologics.

Maternal and fetal alkaline ceramidase 2 is required for placental vascular integrity in mice.

Sphingolipids have been implicated in mammalian placental development and function, but their regulation in the placenta remains unclear. Herein we report that alkaline ceramidase 2 (ACER2) plays a key role in sustaining the integrity of the placental vasculature by regulating the homeostasis of sphingolipids in mice. The mouse alkaline ceramidase 2 gene (Acer2) is highly expressed in the placenta between embryonic day (E) 9.

CD70 Inversely Regulates Regulatory T Cells and Invariant NKT Cells and Modulates Type 1 Diabetes in NOD Mice.

The CD27-CD70 costimulatory pathway is essential for the full activation of T cells, but some studies show that blocking this pathway exacerbates certain autoimmune disorders. In this study, we report on the impact of CD27-CD70 signaling on disease progression in the NOD mouse model of type 1 diabetes (T1D). Specifically, our data demonstrate that CD70 ablation alters thymocyte selection and increases circulating T cell levels.

Genes adapt to outsmart gene-targeting strategies in mutant mouse strains by skipping exons to reinitiate transcription and translation.

Background: Gene disruption in mouse embryonic stem cells or zygotes is a conventional genetics approach to identify gene function in vivo. However, because different gene disruption strategies use different mechanisms to disrupt genes, the strategies can result in diverse phenotypes in the resulting mouse model. To determine whether different gene disruption strategies affect the phenotype of resulting mutant mice, we characterized Rhbdf1 mouse mutant strains generated by three commonly used strategies-definitive-null, targeted knockout (KO)-first, and CRISPR/Cas9.

Loss of TRP53 (p53) accelerates tumorigenesis and changes the tumor spectrum of SJL/J mice.

Known as the guardian of the genome, transformation-related protein 53 (TRP53) is a well -known tumor suppressor. Here, we describe a novel TRP53 deficient mouse model on a tumor prone background-SJL/J mice. The absence of TRP53 (TRP53 nullizygosity) leads to a shift in the tumor spectrum from a non-Hodgkin's-like disease to thymic lymphomas and testicular teratomas at a very rapid tumor onset averaging ~12 weeks of age.

Improved mouse models and advanced genetic and genomic technologies for the study of neutrophils.

Mice have been excellent surrogates for studying neutrophil biology and, furthermore, murine models of human disease have provided fundamental insights into the roles of human neutrophils in innate immunity. The emergence of novel humanized mice and high-diversity mouse populations offers the research community innovative and powerful platforms for better understanding, respectively, the mechanisms by which human neutrophils drive pathogenicity, and how genetic differences underpin the variation in neutrophil biology observed among humans. Here, we review key examples of these new resources.

Identification of as a First Susceptibility Gene for Lithium-Induced Nephrogenic Diabetes Insipidus in Mice.

Background: Lithium, mainstay treatment for bipolar disorder, causes nephrogenic diabetes insipidus and hypercalcemia in about 20% and 10% of patients, respectively, and may lead to acidosis. These adverse effects develop in only a subset of patients treated with lithium, suggesting genetic factors play a role.

Methods: To identify susceptibility genes for lithium-induced adverse effects, we performed a genome-wide association study in mice, which develop such effects faster than humans.

Quantitative Trait Locus and Integrative Genomics Revealed Candidate Modifier Genes for Ectopic Mineralization in Mouse Models of Pseudoxanthoma Elasticum.

Pseudoxanthoma elasticum, a prototype of heritable multisystem ectopic mineralization disorders, is caused by mutations in the ABCC6 gene encoding a putative efflux transporter, ABCC6. The phenotypic spectrum of pseudoxanthoma elasticum varies, and the correlation between genotype and phenotype has not been established. To identify genetic modifiers, we performed quantitative trait locus analysis in inbred mouse strains that carry the same hypomorphic allele in Abcc6 yet with highly variable ectopic mineralization phenotypes of pseudoxanthoma elasticum.

Lack of acute xenogeneic graft- versus-host disease, but retention of T-cell function following engraftment of human peripheral blood mononuclear cells in NSG mice deficient in MHC class I and II expression.

Immunodeficient mice engrafted with human peripheral blood mononuclear cells (PBMCs) support preclinical studies of human pathogens, allograft rejection, and human T-cell function. However, a major limitation of PBMC engraftment is development of acute xenogeneic graft- versus-host disease (GVHD) due to human T-cell recognition of murine major histocompatibility complex (MHC). To address this, we created 2 NOD- scid IL-2 receptor subunit γ ( IL2rg) (NSG) strains that lack murine MHC class I and II [NSG-β-2-microglobulin ( B2M) ( IA IE) and NSG -( K D) ( IA)].

Spontaneous Posterior Segment Vascular Disease Phenotype of a Mouse Model, rnv3, Is Dependent on the Crb1rd8 Allele.

Purpose: To determine the molecular basis of lesion development in a murine model of spontaneous retinal vascularization, rnv3 (retinal vascularization 3, aka JR5558).

Methods: Disease progression of rnv3 was examined in longitudinal studies by clinical evaluation, electroretinography (ERG) and light microscopy analyses. The chromosomal position for the recessive rnv3 mutation was determined by DNA pooling and genome-wide linkage analysis.

RHBDF2-Regulated Growth Factor Signaling in a Rare Human Disease, Tylosis With Esophageal Cancer: What Can We Learn From Murine Models?

Tylosis with esophageal cancer syndrome (TOC) is a rare autosomal dominant proliferative skin disease caused by missense mutations in the rhomboid 5 homolog 2 (RHBDF2) gene. TOC is characterized by thickening of the skin in the palms and feet and is strongly linked with the development of esophageal squamous cell carcinoma. Murine models of human diseases have been valuable tools for investigating the underlying genetic and molecular mechanisms of a broad range of diseases.

Preclinical assessment of antiviral combination therapy in a genetically humanized mouse model for hepatitis delta virus infection.

Chronic delta hepatitis, caused by hepatitis delta virus (HDV), is the most severe form of viral hepatitis, affecting at least 20 million hepatitis B virus (HBV)-infected patients worldwide. HDV/HBV co- or superinfections are major drivers for hepatocarcinogenesis. Antiviral treatments exist only for HBV and can only suppress but not cure infection.

Alkaline ceramidase 2 is essential for the homeostasis of plasma sphingoid bases and their phosphates.

Sphingosine-1-phosphate (S1P) plays important roles in cardiovascular development and immunity. S1P is abundant in plasma because erythrocytes-the major source of S1P-lack any S1P-degrading activity; however, much remains unclear about the source of the plasma S1P precursor, sphingosine (SPH), derived mainly from the hydrolysis of ceramides by the action of ceramidases that are encoded by 5 distinct genes, acid ceramidase 1 ( ASAH1)/ Asah1, ASAH2/ Asah2, alkaline ceramidase 1 ( ACER1)/ Acer1, ACER2/ Acer2, and ACER3/ Acer3, in humans/mice. Previous studies have reported that knocking out Asah1 or Asah2 failed to reduce plasma SPH and S1P levels in mice.

Deletion of a Long-Range Enhancer Disrupts Inner Ear Development in Mice.

Distal enhancers are thought to play important roles in the spatiotemporal regulation of gene expression during embryonic development, but few predicted enhancer elements have been shown to affect transcription of their endogenous genes or to alter phenotypes when disrupted. Here, we demonstrate that a 123.6-kb deletion within the mouse gene is associated with reduced transcription of , a gene located 660 kb away.

Genetic deletion of amphiregulin restores the normal skin phenotype in a mouse model of the human skin disease tylosis.

In humans, gain-of-function (GOF) mutations in RHBDF2 cause the skin disease tylosis. We generated a mouse model of human tylosis and show that GOF mutations in RHBDF2 cause tylosis by enhancing the amount of amphiregulin (AREG) secretion. Furthermore, we show that genetic disruption of AREG ameliorates skin pathology in mice carrying the human tylosis disease mutation.