A mouse model to dissect progesterone signaling in the female reproductive tract and mammary gland

Considering the regulatory complexities of progesterone receptor (PR) action throughout the female reproductive axis and mammary gland, we generated a mouse model that enables conditional ablation of PR function in a spatiotemporal specific manner. Exon 2 of the murine PR gene was floxed to generate a conditional PR allele (PR^flox) in mice. Crossing the PR^flox/flox mouse with the ZP3‐cre transgenic demonstrated that the PR^flox allele recombines to a PR null allele (PR^d). Mice homozygous for the recombined null PR allele (PR^d/d) exhibit uterine, ovarian, and mammary gland defects that phenocopy those of our previously described PR knockout (PRKO) model. Therefore, this conditional mouse model for PR ablation represents an invaluable resource with which to further define in a developmental and/or reproductive stage‐specific manner the individual and integrative roles of distinct PR populations resident in multiple progesterone‐responsive target sites. genesis 48:106–113, 2010. © 2009 Wiley‐Liss, Inc.     

Generation of mice with a conditional null allele of the Jagged2 gene

The Notch signaling pathway is an evolutionarily‐conserved intercellular signaling mechanism, and mutations in its components disrupt embryonic development in many organisms and cause inherited diseases in humans. The Jagged2 (Jag2) gene, which encodes a ligand for Notch pathway receptors, is required for craniofacial, limb, and T cell development. Mice homozygous for a Jag2 null allele die at birth from cleft palate, precluding study of Jag2 function in postnatal and adult mice. We have generated a Jag2 conditional null allele by flanking the first two exons of the Jag2 gene with loxP sites. Cre‐mediated deletion of the Jag2^flox allele generates the Jag2^del2 allele, which behaves genetically as a Jag2 null allele. This Jag2 conditional null allele will enable investigation of Jag2 function in a variety of tissue‐specific contexts. genesis 48:390–393, 2010. © 2010 Wiley‐Liss, Inc.     

Generation of mice with a conditional allele for G6pc

Glucose‐6‐phosphatase‐α (G6Pase‐α or G6PC) catalyzes the hydrolysis of glucose‐6‐phosphate to glucose and is a key enzyme in interprandial glucose homeostasis. Mutations in the human G6PC gene, expressed primarily in the liver, kidney, and intestine, cause glycogen storage disease Type Ia (GSD‐Ia), an autosomal recessive disorder characterized by a disturbed glucose homeostasis. For better understanding of the roles of G6Pase‐α in different tissues and in pathological conditions, we have generated mice harboring a conditional null allele for G6pc by flanking Exon 3 of the G6pc gene with loxP sites. We confirmed the null phenotype by using the EIIa‐Cre transgenic approach to generate mice lacking Exon 3 of the G6pc gene. The resulting homozygous Cre‐recombined null mice manifest a phenotype mimicking G6Pase‐α‐deficient mice and human GSD‐Ia patients. This G6pc conditional null allele will be valuable to examine the consequence of tissue‐specific G6Pase‐α deficiency and the mechanisms of long‐term complications in GSD‐Ia. genesis 47:590–594, 2009. Published 2009 Wiley‐Liss, Inc.     

Generation of a conditional null allele of NADPH oxidase activator 1 (NOXA1)

NADPH oxidase complexes are multiprotein assemblies that generate reactive oxygen species in a variety of mammalian tissues. The canonical phagocytic oxidase consists of a heterodimeric, enzymatic core comprised of the transmembrane proteins, CYBB andCYBA and is regulated, in part, by an “organizing” function of NCF1 and an “activating” activity of NCF2. In contexts outside of the phagocyte, these regulatory functions may be encoded not only by NCF1 and NCF2, but also alternatively by their respective paralogues, NOXO1 and NOXA1. To allow tissue‐specific dissection of Noxa1 function in mouse, we have generated an allele of Noxa1 suitable for conditional inactivation. Moreover, by crossing Noxa1 conditional allele carriers to B6.129S4‐Meox2^tm1(Cre)Sor/J mice, we have generated first, Noxa1‐null heterozygotes, and ultimately, Noxa1‐null homozygotes. Through the thoughtful use of tissue‐specific, Cre‐expressing mouse strains, the Noxa1 conditional allele will offer insight into the roles of NOXA1 in the variety of tissues in which it is expressed. genesis 48:568–575, 2010. © 2010 Wiley‐Liss, Inc.     

Generation of mice with a conditional null allele for Wntless

The Wnt‐signaling pathway is necessary in a variety of developmental processes and has been implicated in numerous pathologies. Wntless (Wls) binds to Wnt proteins and facilitates Wnt sorting and secretion. Conventional deletion of Wls results in early fetal lethality due to defects in body axis establishment. To gain insight into the function of Wls in later stages of development, we have generated a conditional null allele. Homozygous germline deletion of Wls confirmed prenatal lethality and failure of embryonic axis formation. Deletion of Wls using Wnt1‐cre phenocopied Wnt1 null abnormalities in the midbrain and hindbrain. In addition, conditional deletion of Wls in pancreatic precursor cells resulted in pancreatic hypoplasia similar to that previously observed after conditional β‐catenin deletion. This Wls conditional null allele will be valuable in detecting novel Wnt functions in development and disease. genesis 48:554–558, 2010. © 2010 Wiley‐Liss, Inc.     

Gain of function of Tbx1 affects pharyngeal and heart development in the mouse

Mammalian development is highly sensitive to Tbx1 gene dosage reduction. Gene function insights can also be learned from increased or ectopic expression. The authors generated a novel mouse transgenic line, named COET, which expresses Tbx1 upon Cre‐mediated recombination. The authors crossed this transgenic line with Tbx1^Cre animals to activate expression in the Tbx1‐expression domain. Compound mutant COET;Tbx1^Cre/+ animals died after birth and showed heart enlargement. At E18.5, compound mutants showed ventricular septal defects and thymic abnormalities. The authors crossed compound mutants into a Tbx1 null background to understand whether this phenotype is caused by gene overdosage. Results showed that gene dosage reduction at the endogenous locus could not rescue heart and thymic defects, although the transgene rescued the loss of function phenotype. Thus, the transgenic phenotype appears to be due to gain of function. Resultant data demonstrate that Tbx1 expression must be tightly regulated to be compatible with normal embryonic development. genesis 47:188–195, 2009. © 2009 Wiley‐Liss, Inc.     

Generation of a Magoh conditional allele in mice

Magoh encodes a core component of the exon junction complex (EJC), which binds mRNA and regulates mRNA metabolism. Magoh is highly expressed in proliferative tissues during development. EJC components have been implicated in several developmental disorders including TAR syndrome, Richieri–Costa–Pereira syndrome, and intellectual disability. Existing germline null Magoh mice are embryonic lethal as homozygotes and perinatal lethal as heterozygotes, precluding detailed analysis of embryonic and postnatal functions. Here, we report the generation of a new genetic tool to dissect temporal and tissue‐specific roles for Magoh in development and adult homeostasis. This Magoh conditional allele has two loxP sites flanking the second exon. Ubiquitous Cre‐mediated deletion of the floxed allele in a heterozygous mouse (Magoh^del/+) causes 50% reduction of both Magoh mRNA and protein. Magoh^del/+ mice exhibit both microcephaly and hypopigmentation, thus phenocopying germline haploinsufficient Magoh mice. Using Emx1‐Cre, we further show that conditional Magoh deletion in neural progenitors during embryonic development also causes microcephaly. We anticipate this novel conditional allele will be a valuable tool for assessing tissue‐specific roles for Magoh in mammalian development and postnatal processes. genesis 52:752–758, 2014. © 2014 Wiley Periodicals, Inc.     

Generation of mouse conditional and null alleles of the type III sodium‐dependent phosphate cotransporter PiT‐1

Accelerated vascular calcification occurs in several human diseases including diabetes and chronic kidney disease (CKD). In patients with CKD, vascular calcification is highly correlated with elevated serum phosphate levels. In vitro, elevated concentrations of phosphate induced vascular smooth muscle cell matrix mineralization, and the inorganic phosphate transporter‐1 (PiT‐1), was shown to be required. To determine the in vivo role of PiT‐1, mouse conditional and null alleles were generated. Here we show that the conditional allele, PiT‐1^flox, which has loxP sites flanking exons 3 and 4, is homozygous viable. Cre‐mediated recombination resulted in a null allele that is homozygous lethal. Examination of early embryonic development revealed that the PiT‐1^{Δe3,4/Δe3,4} embryos displayed anemia, a defect in yolk sac vasculature, and arrested growth. Thus, conditional and null PiT‐1 mouse alleles have been successfully generated and PiT‐1 has a necessary, nonredundant role in embryonic development. genesis 47:858–863, 2009. © 2009 Wiley‐Liss, Inc.     

Generation of an EphA4 conditional allele in mice

Ephrins and Eph receptor tyrosine kinases are cell‐surface molecules that serve a multitude of functions in cell–cell communication in development, physiology, and disease. EphA4 is a promiscuous member of the EphA subclass of Eph receptors and can bind to both EphrinAs and EphrinBs. In addition to its well‐established roles in guiding the development of neuronal connectivity, EphA4 has been implicated for a role in synaptic plasticity, vascular formation, axon regeneration, and central nervous system repair following injury. However, the study of its role in the adult stage has been hampered by confounding developmental defects in EphA4 germline mutants. Here, we report the generation and molecular characterization of an EphA4 conditional allele along with a novel null allele with a knockin fluorescent reporter gene (mCFP). The conditional allele will be useful in ascertaining postdevelopmental and/or cell type‐specific function of EphA4 in physiology, injury, and disease. genesis 48:101–105, 2010. © 2009 Wiley‐Liss, Inc.     

Important Functional Roles of Basigin in Thymocyte Development and T cell Activation

Basigin is a highly glycosylated transmembrane protein that is expressed in a broad range of tissues and is involved in a number of physiological and pathological processes. However, the in vivo role of basigin remains unknown. To better understand the physiological and pathological functions of basigin in vivo, we generated a conditional null allele by introducing two loxP sites flanking exons 2 and 7 of the basigin gene (Bsg). Bsg^fl/fl mice were born at the expected Mendelian ratio and showed a similar growth rate compared with wildtype mice. After crossing these mice with Lck-Cre transgenic mice, basigin expression was specifically inactivated in T cells in the resulting Lck-Cre; Bsg^fl/fl mice. Although the birth and growth rate of Lck-Cre; Bsg^fl/fl mice were similar to control mice, thymus development was partially arrested in Lck-Cre; Bsg^fl/fl mice, specifically at the CD4⁺CD8⁺ double-positive (DP) and CD4 single-positive (CD4⁺CD8^-, CD4SP) stages. In addition, CD4⁺ T cell activation was enhanced upon Concanavalin A (Con A) or anti-CD3/anti-CD28 stimulation but not upon PMA/Ionomycin stimulation in the absence of basigin. Overall, this study provided the first in vivo evidence for the function of basigin in thymus development. Moreover, the successful generation of the conditional null basigin allele provides a useful tool for the study of distinct physiological or pathological functions of basigin in different tissues at different development stages.