Two spontaneous mutant alleles, rb and rb2J, of the Epha4 gene on mouse Chromosome 1

Two spontaneous mutant alleles, rb and rb^2J, of the Epha4 gene on mouse Chromosome 1

Susan A. Cook, Eva M. Eicher, Richard M. Samples, Roderick T. Bronson and Muriel T. Davisson

Source of Support: NIH/NCRR grant RR01183 (M. Davisson and Eva Eicher, PIs) and NCI Cancer Center Core Grant CA34196

Mutation (allele) symbols: rb and rb^2J

Mutation (allele) names: rabbit and rabbit 2 Jackson

Gene symbol: Epha4

Strain of Origin: C57BL/6J

Current Strain names: C3Sn.B6-Epha4^rb/EiGrsrJ and C57BL/6J-Epha4^rb^2J/EiGrsrJ

Stock #s: 001502 for rb, 003129 for rb^2J(view JAX® Mice Data Sheet for additional information including Price and Supply Information)

Phenotype categories: neurological/behavioral: motor capabilities/coordination/movement anomalies

Origin and Description

Both rb and rb^2J alleles arose on the C57BL/6J background as spontaneous recessive mutations at The Jackson Laboratory in 1964 and 1997, respectively. The rb allele was transferred to the C3H/HeSnJ background and that congenic stock is designated C3Sn.B6-Epha4^rb/EiGrsrJ. Mutants for either allele are visibly identified by 2.5 – 3 weeks of age by a constant hopping gait of the hind limbs; the forelimbs may show a hopping gait or move normally. When picked up by the tail, mutants of either allele clasp their hindlimbs and show reduced ability to hold onto an edge; forelimbs show normal strength. More severely affected, rb^2J/rb^2J mice also lean frequently to either side. rb/rb females breed well, producing 6-8 pups per litter. rb^2J/rb^2J females may deliver 2-3 pups per litter but often exhibit poor nurturing. The rb mutation, currently at N37F1, is maintained by alternating generations of backcrossing female mutants to wildtype males of the C3H/HeSnJ strain and then intercrossing heterzygotes. The rb^2J mutation currently at F200+18 is maintained by mating homozygous female mutants to heterozygous siblings. The C57BL/6J-Epha4^rb2J/EiGrsrJ colony is supplemented by transplantation of mutant ovaries into histocompatible or immuno-deficient hosts.
DNA of both mutations has been cryopreserved in TJL DNA Resource (stock numbers listed above). Embryos carrying the rb allele are cryopreserved as Stock # 000938 (parents are B6J wildtype females x STOCK heterozygous males) and Stock # 001502 (parents are C3Sn.B6 congenic heterozygotes). Embryos carrying the rb^2J allele are cryopreserved as Stock # 003129 (parents are B6J wildtype females x heterozygous males).

Genetic Analysis

The rb2J allele is a remutation at the rb locus. (A C3Sn.B6-rb/rb female mated to a C57BL/6J +/rb2J male produced 4 visibly affected mice from 2 litters of 11 total progeny.) An early linkage cross located rb between the isoenzyme genes ldh1 and Pep3 on mouse Chromosome 1. To refine the position of rb on Chromosome 1, we used MIT microsatellite markers to type 75 F2 mutants from a B6-rb2J x CAST/Ei intercross. The results are given in centimorgans +/- standard error: D1Mit77—2.01 +/- 1.14—[D1Mit181, D1Mit7, D1Mit46, D1Mit333, D1Mit132, rb2J] - 0.67 +/- 0.66—[D1Mit216, D1Mit44]. The order of markers was determined with Map Manager (Manly, 1993) and the linkage data are available from MGD accession number J: 87289). MGSC (2003) as shown by Ensembl (2003) agrees with our mapping order and places the cosegregant D1Mit132 at map position 77.8 Mb near Epha4 (Eph receptor A4) at 78.1 Mb. Because targeted and gene-trap mutations of the Epha4 gene cause a hopping gait in mice, it seemed likely that rb and rb2J are mutations within the Epha4 gene. The Tessier-Lavigne Laboratory at Stanford University kindly provided mice carrying a gene-trap mutation in Epha4 for complementation testing. A Epha4Gt(pGT1TM)38Wcs/+ female was mated to a rb/+ male. Of the 7 total progeny produced, 1 female and 2 males displayed a hopping gait with no leaning and were visibly indistinguishable from rb/rb mutants. Similarly, the reciprocal mating produced 3 mutants (2 females and 1 male) out of 24 total young. These results indicate that rb and rb-2J are mutations within the Epha4 gene.

Pathology

Our standard pathology screen revealed no anomalies in either rb/rb or rb^2J/rb^2J adults. ABR (auditory brainstem response) testing revealed no significant hearing loss in rb/rb mice at 4 weeks of age or in rb^2J/ rb^2Jmice at 4 - 6 months of age. rb/rb mutants on the congenic background do have retinal degeneration characteristic of most C3H strains. rb^2J/rb^2J mutants have normal ERGs (electroretinograms), but may have a retinal histopathology awaiting verification. Fresh sperm specimens from two rb^2J/rb^2J males at 4 months of age appeared normal by light microscopy.

Acknowledgements

We thank Nicole Look for discovery of the phenotypic deviant that carried the rb^2J mutation. We acknowledge Coleen Marden, Heping Yu and Norman Hawes for clinical assistance and Pat Ward-Bailey for Web posting. We’re grateful to Susan Ackerman, Ph.D for predicting Epha4 as the correct candidate gene even prior to our mapping of rb^2J with MIT markers. We especially thank Drs. Marc Tessier-Lavigne and William C. Skarnes for providing mice with a gene-trap mutation in Epha4.

References

MGD 2004, Mouse Genome Database, Mouse Genome Informatics Project, The Jackson Laboratory, Bar Harbor, ME. (URL: http://www.informatics.jax.org).

MGSC 16.30.1, Mouse Genome Sequencing Consortium 2003
(URL: http://www.ensembl.org/Mus_musculus/).

Manley KF (1993) A MacIntosh program for storage and analysis of experimental mapping data. Mamm Genome 4, 303-313.

Last Modified: June 30, 2009

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