Overview

When Myh9^flox mice are crossed with Pod::Cre mice resulting offspring will have exon 1 deleted in kidney podocytes. Double mutant mice on the C57BL/6 background do not develop spontaneous proteinuria or renal insufficiency, however, when subjected to environmental stress via doxyrubicin hydrochloride (Adriamycin), Myh9 podocyte-deleted mice develop albuminuria, focal and segmental glomerulosclerosis, foot process fusion and foot process effacement.

Donating Investigator

Duncan Johnstone, University of Pennsylvania

Genetic overview

Genetic Background	Generation

Tg(NPHS2-cre)295Lbh

Allele Type
Transgenic (Recombinase-expressing)

Myh9^tm1.1Gac

Allele Type	Gene Symbol	Gene Name
Targeted (Conditional ready (e.g. floxed), No functional change)	Myh9	myosin, heavy polypeptide 9, non-muscle

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Research Applications

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Details

Detailed Description

Myh9 (myosin, heavy chain 9, non-muscle) encodes a nonmuscle myosin IIA heavy chain involved in several functions including cytokinesis, cell motility and maintenance of cell shape. Genome wide association studies associate single nucleotide polymorphisms (SNPs) in Myh9 with chronic kidney disease in African-Americans. Myh9^flox mutant mice possess loxP sites flanking exon 1. Homozygous mice are viable and fertile.

The podocin nephrosis 2 homolog (NPHS2) promoter directs expression of Cre recombinase to podocytes in the newborn and adult kidneys. Expression is minimal or undetectable in all other tissues tested and in 8.5 dpc embryos. Mice homozygous for Pod::Cre are viable and fertile.

When Myh9^flox mice are crossed with Pod::Cre mice resulting offspring will have exon 1 deleted in kidney podocytes. Double mutant mice on the C57BL/6 background do not develop spontaneous proteinuria or renal insufficiency, however, when subjected to environmental stress via doxyrubicin hydrochloride (Adriamycin), Myh9 podocyte-deleted mice develop albuminuria, focal and segmental glomerulosclerosis, foot process fusion and foot process effacement.

This strain is unpublished on the FVB background. The donating investigator indicates that FVB mice are more sensitive to glomerular disease than C57BL/6 mice.

Development

The Myh9^tm1.1Gac targeting vector was designed to insert a loxP site upstream of exon 1 followed by a loxP-flanked neomycin resistance (neo) cassette downstream of exon 1 in the Myh9 gene. The construct was electroporated into 129/Sv-derived embryonic stem (ES) cells. Transient Cre expression in targeted cells excised the neo cassette leaving exon 1 flanked by loxP sites. Correctly targeted ES cells were injected into blastocysts. The resulting chimeric animals were crossed to C57BL/6 mice for at least 5 generations.

The Pod::Cre (or 2.5P-Cre) transgene was designed with 2.5 kb fragment of the human podocin nephrosis 2 homolog (NPHS2) promoter/enhancer region followed by a Cre recombinase cassette. The transgene was microinjected into the pronuclei of fertilized oocytes from (C57BL/6 x SJL)F2 mice. Founder line 295 was established and used to generate mutant mice. The resulting transgenic mice were subsequently bred to wild-type C57BL/6 mice for an unknown number of generations.

Both strains were transferred to the University of Pennsylvania and intercrossed. Double mutant mice (Pod::Cre;Myh9f) were backcrossed to FVB/NJ mice by marker assisted selection (speed congenic) for 4 generations and then by traditional backcross for 10 generations.
The donating investigator maintains the strain by Pod::Cre/+ Myh9f/+ x FVB/NJ or Myh9f/f. Upon arrival, mice were bred to FVB/NJ for at least 1 generation to establish the colony.

Expression Data

Expressed Gene	cre, cre recombinase, bacteriophage P1
Site of Expression	cre recombinase expression in podocytes within kidney glomeruli.
Site of Expression

Control Suggestions

001800 FVB/NJ

Additional Information

Considerations for Choosing Controls

Selected References

Johnstone DB; Zhang J; George B; Leon C; Gachet C; Wong H; Parekh R; Holzman LB. 2011. Podocyte-specific deletion of Myh9 encoding nonmuscle myosin heavy chain 2A predisposes mice to glomerulopathy. Mol Cell Biol 31(10):2162-70PubMed: 21402784 MGI: J:173971

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Genetics

Tg(NPHS2-cre)295Lbh

Allele Symbol: Tg(NPHS2-cre)295Lbh

Allele Name	transgene insertion 295, Lawrence B Holzman
Allele Type	Transgenic (Recombinase-expressing)
Allele Synonym(s)	2.5P-Cre; NPHS2.Cre; pod-Cre; podocin-Cre; PodoCre; Tg(NPHS2-cre)1Lbh
Gene Symbol and Name	Tg(NPHS2-cre)295Lbh, transgene insertion 295, Lawrence B Holzman
Gene Synonym(s)
Promoter	NPHS2, NPHS2, podocin, human
Expressed Gene	cre, cre recombinase, bacteriophage P1
Site of Expression	cre recombinase expression in podocytes within kidney glomeruli.
Strain of Origin	(C57BL/6 x SJL)F2
Chromosome	UN
General Note	24 transgenic founders were created. Phenotypic analysis was performed on founder line #295.
Molecular Note	The human NPHS2 promoter, including the entire 5' untranslated region, was used to drive cre recombinase expression in podocytes.
Mutations Made By	Lawrence Holzman, University of Pennsylvania

Myh9^tm1.1Gac

Allele Symbol: Myh9^tm1.1Gac

Allele Name	targeted mutation 1.1, Christian Gachet
Allele Type	Targeted (Conditional ready (e.g. floxed), No functional change)
Allele Synonym(s)	MYH9^Flx; Myh9flox
Gene Symbol and Name	Myh9, myosin, heavy polypeptide 9, non-muscle
Gene Synonym(s)
Strain of Origin	129
Chromosome	15
Molecular Note	A loxP site was inserted upstream of exon 1, and a floxed neo cassette was inserted downstream of exon 1. Transient expression in ES cells removed the neo cassette leaving exon 1 floxed.

Disease/Phenotype

Disease Terms

Research Areas By Phenotype

This mouse can be used to support research in many areas including:

Mammalian Phenotype Terms by Genotype

The following phenotype information is associated with a similar, but not exact match to this JAX^® Mice strain

Genotype: Myh9^tm1.1Gac/Myh9^tm1.1Gac Tg(NPHS2-cre)295Lbh/0
involves: 129 * C57BL/6 * SJL

renal/urinary system phenotype

fused podocyte foot processes
- after adriamycin injection, mice exhibit patches of foot process fusion, unlike similarly-treated control mice

glomerulosclerosis
- ic glomeruli
- at 6 weeks after adriamycin injection, mice develop focal and segmental glomerulosclerosis ranging from mild segmental sclerosis to severe global sclerosis, unlike similarly-treated control mice which develop foci of mild sclerosis but no globally sclerotic glomeruli

increased renal glomerulus basement membrane thickness
- after adriamycin injection, mice exhibit thickening of the basement membrane in some glomerular loops, unlike similarly-treated control mice

podocyte microvillus transformation
- after adriamycin injection, mice exhibit evidence of pseudovillous transformation, unlike similarly-treated control mice

renal cast
- at 6 weeks after adriamycin injection, mice exhibit numerous proteinaceous casts, unlike similarly-treated control mice

albuminuria
- at 3, 4, and 6 weeks after adriamycin injection, mice develop significantly more albuminuria than similarly-treated control mice

podocyte foot process effacement
- after adriamycin injection, mice exhibit patches of severe foot process effacement, unlike similarly-treated control mice

homeostasis/metabolism phenotype

increased susceptibility to injury
- following injection with doxorubicin hydrochloride (adriamycin; 15 ug/g) to induce kidney podocyte injury, 9-11 month old mice develop severe proteinuria and glomerulosclerosis, unlike similarly-treated control mice

albuminuria
- at 3, 4, and 6 weeks after adriamycin injection, mice develop significantly more albuminuria than similarly-treated control mice

mammalian phenotype

homeostasis/metabolism phenotype
- Normal - normal urine albumin/creatinine ratios at 3-9 months of age relative to control mice
- Normal - normal serum creatinine and BUN levels at 9 months of age relative to control mice
- Normal - no excessive proteinuria at 1-9 months of age relative to control mice

cardiovascular system phenotype
- Normal - no significant difference in tail cuff blood pressure at 11-12 months of age relative to control mice

growth/size/body region phenotype
- Normal - normal body weight at 9 months of age relative to control mice

renal/urinary system phenotype
- Normal - normal urine albumin/creatinine ratios at 3-9 months of age relative to control mice
- Normal - surprisingly, mice are viable and do not develop overt chronic kidney disease up to 9 months of age
- Normal - no excessive proteinuria at 1-9 months of age relative to control mice

References

Johnstone DB; Zhang J; George B; Leon C; Gachet C; Wong H; Parekh R; Holzman LB. 2011. Podocyte-specific deletion of Myh9 encoding nonmuscle myosin heavy chain 2A predisposes mice to glomerulopathy. Mol Cell Biol 31(10):2162-70PubMed: 21402784 MGI: J:173971

Additional - Myh9^tm1.1Gac related

Additional - Tg(NPHS2-cre)295Lbh related

Technical Support

CONTACT TECHNICAL SUPPORT

Genotyping Protocols
- Standard PCR:Myh9
- Genotyping resources and troubleshooting
Breeding Considerations

While maintaining a live colony, these mice are bred as double homozygotes.
- Additional Breeding and Husbandry Support
Citation
When using the FVB.Cg-Myh9^tm1.1Gac Tg(NPHS2-cre)295Lbh/Mmjax mouse strain in a publication, please cite the originating article(s) and include MMRRC stock #36742 in your Materials and Methods section.

Animal Health Reports

Facility Barrier Level Descriptions

Production of mice from cryopreserved embryos or sperm occurs in a maximum barrier room, G200

Terms Of Use

General Terms and Conditions

See MMRRC for Additional Conditions of Distribution

Questions about Terms of Use

Licensing Information

Phone: 207-288-6470

Email: TechTran@jax.org

Donating Investigator

Genetic overview

Research Applications

Detailed Description

Development

Expression Data

Control Suggestions

Additional Information

Selected References

Tg(NPHS2-cre)295Lbh

Allele Symbol: Tg(NPHS2-cre)295Lbh

Myh9tm1.1Gac

Allele Symbol: Myh9tm1.1Gac

Disease Terms

Research Areas By Phenotype

This mouse can be used to support research in many areas including:

Mammalian Phenotype Terms by Genotype

Genotype: Myh9tm1.1Gac/Myh9tm1.1Gac Tg(NPHS2-cre)295Lbh/0involves: 129 * C57BL/6 * SJL

renal/urinary system phenotype

homeostasis/metabolism phenotype

mammalian phenotype

References

Additional - Myh9tm1.1Gac related

Additional - Tg(NPHS2-cre)295Lbh related

Genotyping Protocols

Breeding Considerations

Citation

Animal Health Reports

Production of mice from cryopreserved embryos or sperm occurs in a maximum barrier room, G200

The Jackson Laboratory's Genotype Promise

Terms Of Use

Licensing Information

Myh9^tm1.1Gac

Allele Symbol: Myh9^tm1.1Gac

Genotype: Myh9^tm1.1Gac/Myh9^tm1.1Gac Tg(NPHS2-cre)295Lbh/0
involves: 129 * C57BL/6 * SJL

Additional - Myh9^tm1.1Gac related