A physical map of the genomic region on mouse chromosome 3 containing the hindshaker (hsh) mutation

Hindshaker (hsh), a spontaneous, autosomal recessive mouse mutation, displays a developmentally dependent tremor of the hindquarters due to hypomyelination in the CNS. This myelin deficit is followed by progressive, but incomplete, recovery by postnatal day 42. Herein we describe the construction of a genomic contig spanning the interval between the markers D3Mit187 (42.4 cM) and D3Mit232 (45.2 cM) on mouse chromosome 3, which we have previously shown to contain the hsh mutation. A physical map, covering approximately 3.5 Mb, was constructed from a series of overlapping yeast and bacterial artificial chromosomes. A 1.2- to 1.4-Mb segment central to the contig was compared extensively with the syntenic regions in human (chromosome 1q21-q23) and rat (chromosome 2). We present new data on 10 genes erroneously assigned to this area and on another 6 genes previously assigned elsewhere. For absent genes, our work suggests that they are telomeric to the region encompassed in our map. Accordingly, our findings both map the area surrounding the hsh mutation and present important corrections to the current maps in an area rich in genes related to the nervous system.     

Effects of Rumpshaker Mutation on CNS Myelin Composition and Structure

Abstract: Myelinated CNS tissues from homozygous/hemizygous and heterozygous jimpy rumpshaker jp ^rsh mutant mice were examined to determine the consequences on myelin structure of this mutation in the proteolipid protein (PLP) gene. Polyacrylamide gel electrophoresis and immunoblotting of brain homogenates confirmed that there was a decrease in PLP levels on the B6C3 genetic background onto which this gene was bred. We also observed an increase in level of a protein band that could correspond to the uncharacterized 10-kDa PLP previously reported in jp ^rsh mice on an Rb(1.3) 1Bnr background. High-performance TLC and densitometry of lipids from brain homogenate and isolated myelin revealed a decrease in content of cerebrosides and sulfatides. Electron microscopy on optic nerves revealed that normal radial component is retained in jp ^rsh myelin, further substantiating that PLP is not a component of this junctional complex. X-raydiffraction measurements on unfixed optic nerves showed that the jp ^rsh period is 5–10 Å larger than normal. Moreover, jp ^rsh optic nerve myelin was unstable, as evidenced by a continual increase in the period postdissection. jp ^rsh myelin that was equilibrated at varying pH and ionic strength typically had a larger than normal period under all conditions (both swelling and compacting). Our findings thus demonstrate that the biochemical abnormalities in the jp ^rsh mutant correlate with a wider periodicity and less stable packing of the myelin.     

Mapping of the dysmyelinating murine Hindshaker mutation to a 1.2-cM interval on chromosome 3

Hindshaker (hsh) is a novel, spontaneous, autosomal recessive mouse mutation displaying a myelin deficit, predominantly in the spinal cord. It is characterized by developmentally dependent hypomyelination, first evident at postnatal day (P) 10, followed by progressive but incomplete recovery by P42. Hypomyelination is associated with a decreased number of mature oligodendrocytes, which fail to form complete myelin sheaths. Heterozygotes are phenotypically normal, and the hsh mutation shows considerable variation in penetrance and expression depending on genetic background, indicating the influence of modifying loci. Here, we followed an outcross/backcross breeding strategy in conjunction with genotyping for microsatellites and a novel marker for the gene S100a4. We describe the genomic mapping of the hsh mutation to within a 1.2-cM region near the centromere of mouse chromosome 3. We found that hsh is flanked between D3Mit187 proximally and S100a4 distally. The area containing hsh is gene-rich, with a high proportion of the genes specific to nervous tissue. Identification of the hsh mutation will aid our understanding of processes important in regional control of oligodendrocyte development and myelination.     

The early phenotype associated with the jimpy mutation of the proteolipid protein gene

The jimpy mutation of the X-linked proteolipid protein (Plp) gene causes dysmyelination and premature death of the mice. The established phenotype is characterised by severe hypomyelination, increased numbers of dead oligodendrocytes and astrocytosis. The purpose of this study was to define the earliest cellular abnormalities in the cervical spinal cord. We find that on the first and third postnatal days the amount of myelin in jimpy spinal cord is approximately 20% of wild-type. However, the total glial cell density, the number of dead glial cells and the number and distribution of Plp-positive cells, as assessed by in situ hybridization, are similar to wild-type during the first week of life. Immunostaining of cryosections has identified that jimpy spinal cords express on schedule, a variety of antigens associated with mature oligodendrocytes. Dissociated oligodendrocytes, cultured for 18 hours to reflect their in vivo differentiation, express MBP and surface myelin-associated glycoprotein at the same frequency as wild-type. By comparison, the proportion of jimpy oligodendrocytes expressing surface myelin/oligodendrocyte glycoprotein is reduced by approximately 34%. In vivo, however, only a small minority of axons is surrounded by a collar of myelin-associated glycoprotein, suggesting that the majority of jimpy oligodendrocytes fail to make appropriate ensheathment of axons. Although the DM20 isoform is expressed in the embryonic CNS prior to myelin formation, the cellular abnormalities appear to correspond to the time at which the Plp isoform becomes predominant. The results suggest that the primary abnormality in jimpy is the inability of oligodendrocytes to properly associate with, and then ensheath, axons and that oligodendrocyte death compounds, rather than initiates, the established phenotype.     

A Novel Gelsolin Isoform Expressed by Oligodendrocytes in the Central Nervous System

Abstract: Two isoforms of the Ca²⁺-sensitive, actin-binding protein gelsolin have been identified thus far; one is an intracellular protein, cytoplasmic gelsolin, and the other is a secretory protein called plasma gelsolin. Gelsolin expression in the mammalian CNS appears to be localized mainly to oligodendrocytes where it is presumed that the cytoplasmic isoform predominates. Here, we show that oligodendrocytes not only contain cytoplasmic gelsolin, but they also express a novel gelsolin isoform that we have named gelsolin-3. Cytoplasmic gelsolin, plasma gelsolin, and gelsolin-3 arise by alternative splicing from the same gene. The N-terminal amino acid sequence unique to gelsolin-3 is shown to be encoded by a single exon in a region previously thought to be an intron in the human gelsolin gene. In situ hybridization analysis confirmed that gelsolin-3 mRNA is localized primarily to oligodendrocytes in rat brain. In other tissues, gelsolin-3 shows a more restricted pattern of expression than cytoplasmic gelsolin. These data support the view that the gelsolin isoforms have differential roles in the regulation of the actin cytoskeleton.