Loss-of-function mutations of FIG4 are responsible for neurological disorders in human and mouse that result from reduced abundance of the signaling lipid PI(3,5)P2. In contrast, loss-of-function mutations of the phosphoinositide kinase PIP4K2C result in elevated abundance of PI(3,5)P2. These opposing effects on PI(3,5)P2 suggested that we might be able to compensate for deficiency of FIG4 by reducing expression of PIP4K2C. To test this hypothesis in a whole animal model, we generated triallelic mice with genotype Fig 4-/-, Pip4k2c+/-; these mice are null for Fig 4 and haploinsufficient for Pip4k2c. The neonatal lethality of Fig 4 null mice in the C57BL/6J strain background was rescued by reduced expression of Pip4k2c. The lysosome enlargement characteristic of Fig 4 null cells was also reduced by heterozygous loss of Pip4k2c. The data demonstrate interaction between these two genes, and suggest that inhibition of the kinase PIPK4C2 could be a target for treatment of FIG4 deficiency disorders such as Charcot-Marie-Tooth Type 4J and Yunis-Varón Syndrome.
The Rbfox proteins (Rbfox1, Rbfox2, and Rbfox3) regulate the alternative splicing of many important neuronal transcripts and have been implicated in a variety of neurological disorders. However, their roles in brain development and function are not well understood, in part due to redundancy in their activities. Here we show that, unlike Rbfox1 deletion, the CNS-specific deletion of Rbfox2 disrupts cerebellar development. Genome-wide analysis of Rbfox2(-/-) brain RNA identifies numerous splicing changes altering proteins important both for brain development and mature neuronal function. To separate developmental defects from alterations in the physiology of mature cells, Rbfox1 and Rbfox2 were deleted from mature Purkinje cells, resulting in highly irregular firing. Notably, the Scn8a mRNA encoding the Na(v)1.6 sodium channel, a key mediator of Purkinje cell pacemaking, is improperly spliced in RbFox2 and Rbfox1 mutant brains, leading to highly reduced protein expression. Thus, Rbfox2 protein controls a post-transcriptional program required for proper brain development. Rbfox2 is subsequently required with Rbfox1 to maintain mature neuronal physiology, specifically Purkinje cell pacemaking, through their shared control of sodium channel transcript splicing.
We previously reported that autosomal recessive demyelinating Charcot-Marie-Tooth (CMT) type 4B1 neuropathy with myelin outfoldings is caused by loss of MTMR2 (Myotubularin-related 2) in humans, and we created a faithful mouse model of the disease. MTMR2 dephosphorylates both PtdIns3P and PtdIns(3,5)P2, thereby regulating membrane trafficking. However, the function of MTMR2 and the role of the MTMR2 phospholipid phosphatase activity in vivo in the nerve still remain to be assessed. Mutations in FIG4 are associated with CMT4J neuropathy characterized by both axonal and myelin damage in peripheral nerve. Loss of Fig4 function in the plt (pale tremor) mouse produces spongiform degeneration of the brain and peripheral neuropathy. Since FIG4 has a role in generation of PtdIns(3,5)P2 and MTMR2 catalyzes its dephosphorylation, these two phosphatases might be expected to have opposite effects in the control of PtdIns(3,5)P2 homeostasis and their mutations might have compensatory effects in vivo. To explore the role of the MTMR2 phospholipid phosphatase activity in vivo, we generated and characterized the Mtmr2/Fig4 double null mutant mice. Here we provide strong evidence that Mtmr2 and Fig4 functionally interact in both Schwann cells and neurons, and we reveal for the first time a role of Mtmr2 in neurons in vivo. Our results also suggest that imbalance of PtdIns(3,5)P2 is at the basis of altered longitudinal myelin growth and of myelin outfolding formation. Reduction of Fig4 by null heterozygosity and downregulation of PIKfyve both rescue Mtmr2-null myelin outfoldings in vivo and in vitro.
Abstract The original article to which this Erratum refers was published in Developmental Dynamics 233:368–372 Developmental Dynamics(2005) 233(2) 368–372
The mnd2 mutation on mouse chromosome 6 produces a progressive neuromuscular disorder. To determine the gene content of the 400-kb mnd2 nonrecombinant region, we sequenced 108 kb of mouse genomic DNA and 92 kb of human genomic sequence from the corresponding region of chromosome 2p13.3. Three genes with the indicated sizes and intergenic distances were identified: D6Mm5e (>/=81 kb)-787 bp-DOK (2 kb)-845 bp-LOR2 (>/=6 kb). D6Mm5e is expressed in many tissues at very low abundance and the predicted 526-residue protein contains no known functional domains. DOK encodes the p62(dok) rasGAP binding protein involved in signal transduction. LOR2 encodes a novel lysyl oxidase-related protein of 757 amino acid residues. We describe a simple search protocol for identification of conserved internal exons in genomic sequence. Evolutionary conservation proved to be a useful criterion for distinguishing between authentic exons and artifactual products obtained by exon amplification, RT-PCR, and 5' RACE. Conserved noncoding sequence elements longer than 80 bp with >/=75% nucleotide sequence identity comprise approximately 1% of the genomic sequence in this region. Comparative analysis of this human and mouse genomic DNA sequence was an efficient method for gene identification and is independent of developmental stage or quantitative level of gene expression. [The sequence data described in this paper have been submitted to the GenBank data library under the following accession numbers: AC003061, mouse BAC clone 245c12; AC003065, human BAC clone h173(E10); AF053368, mouse Lor2 cDNA; AF084363, 108-kb contig from mouse BAC 245c12; AF084364, mouse D6Mm5e cDNA.]
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