Myelin Proteolipid Protein Gene Research Articles

Plp1 in the enteric nervous system is preferentially expressed during early postnatal development in mouse as DM20, whose expression appears reliant on an intronic enhancer.

Recently, the myelin proteolipid protein gene (Plp1) was shown to be expressed in the glia of the enteric nervous system (ENS) in mouse. However, beyond this, not much is known about its expression in the intestine. To address this matter, we investigated Plp1 expression at the mRNA and protein levels in the intestine of mice at different ages (postnatal days 2, 9, 21, and 88). In this study, we show that Plp1 expression preferentially occurs during early postnatal development, primarily as the DM20 isoform. Western blot analysis indicated that DM20 migrated according to its formula weight when isolated from the intestine. However, mobilities of both PLP and DM20 were faster than expected when procured from the brain. The 6.2hPLP(+)Z/FL transgene, which uses the first half of the human PLP1 gene to drive expression of a lacZ reporter gene, recapitulated the developmental pattern observed with the native gene in the intestine, indicating that it can be used as a proxy for Plp1 gene expression. As such, the relative levels of β-galactosidase (β-gal) activity emanating from the 6.2hPLP(+)Z/FL transgene suggest that Plp1 expression is highest in the duodenum, and decreases successively along the segments, toward the colon. Moreover, removal of the wmN1 enhancer region from the transgene (located within Plp1 intron 1) resulted in a dramatic reduction in both transgene mRNA levels and β-gal activity in the intestine, throughout development, suggesting that this region contains a regulatory element crucial for Plp1 expression. This is consistent with earlier studies in both the central and peripheral nervous systems, indicating that it may be a common (if not universal) means by which Plp1 gene expression is governed.

PLP1-lacZ transgenic mice reveal that splice variants containing "human-specific" exons are relatively minor in comparison to the archetypal transcript and that an upstream regulatory element bolsters expression during early postnatal brain development.

Much of what is known about the mechanisms that control the developmental expression of the myelin proteolipid protein gene (PLP1) has been attained through use of transgenic animal models. In this study, we analyzed expression of related transgenes which utilize PLP1 genomic DNA from either human or mouse to drive expression of a lacZ reporter. Human PLP1 (hPLP1) sequence span either the proximal 6.2 or 2.7 kb of 5'-flanking DNA to an internal site in Exon 2, while those from mouse comprise the proximal 2.3 kb of 5'-flanking DNA to an analogous site in Exon 2. Transgenes with hPLP1 sequence were named, in part, to the amount of upstream sequence they have [6.2hPLP(+)Z/FL and 2.7hPLP(+)Z]. The transgene containing mouse sequence is referred to here as mPLP(+)Z, to denote the species origin of PLP1 DNA. Mice which harbor the 6.2hPLP(+)Z/FL transgene were used as a model system to investigate the developmental expression of splice variants that incorporate supplementary exons from what is classically defined as PLP1 intron 1. While expression of the splice variants were detected in brain through RT-PCR analysis, they are present at much lower levels relative to the archetypal (classic) transcript. Additionally, we show that mice which harbor the 6.2hPLP(+)Z/FL transgene demonstrate wide-ranging expression throughout brain at P2, whereas expression of mPLP(+)Z is quite limited at this age. Therefore, we generated new transgenic mouse lines with the 2.7hPLP(+)Z transgene, which contains hPLP1 sequence orthologous to just that in mPLP(+)Z. Of the seven lines analyzed, six showed higher levels of 2.7hPLP(+)Z expression in brain at P21 compared to P2; the other line expressed the transgene, only weakly, at either age. This trend, coupled with the robust expression observed for 6.2hPLP(+)Z/FL at P2, suggests that the distal 3.5 kb of 5'-flanking PLP1 DNA specific to 6.2hPLP(+)Z/FL contains regulatory element(s) important for promoting early postnatal expression in brain.

Ethanol modulation of hippocampal neuroinflammation, myelination, and neurodevelopment in a postnatal mouse model of fetal alcohol spectrum disorders

Fetal alcohol spectrum disorders (FASD) are alarmingly common and result in significant personal and societal loss. Neuropathology of the hippocampus is common in FASD leading to aberrant cognitive function. In the current study, we evaluated the effects of ethanol on the expression of a targeted set of molecules involved in neuroinflammation, myelination, neurotransmission, and neuron function in the developing hippocampus in a postnatal model of FASD. Mice were treated with ethanol from P4-P9, hippocampi were isolated 24 h after the final treatment at P10, and mRNA levels were quantitated by qRT-PCR. We evaluated the effects of ethanol on both pro-inflammatory and anti-inflammatory molecules in the hippocampus and identified novel mechanisms by which ethanol induces neuroinflammation. We further demonstrated that ethanol decreased expression of molecules associated with mature oligodendrocytes and greatly diminished expression of a lacZ reporter driven by the first half of the myelin proteolipid protein (PLP) gene (PLP1). In addition, ethanol caused a decrease in genes expressed in oligodendrocyte progenitor cells (OPCs). Together, these studies suggest ethanol may modulate pathogenesis in the developing hippocampus through effects on cells of the oligodendrocyte lineage, resulting in altered oligodendrogenesis and myelination. We also observed differential expression of molecules important in synaptic plasticity, neurogenesis, and neurotransmission. Collectively, the molecules evaluated in these studies may play a role in ethanol-induced pathology in the developing hippocampus and contribute to cognitive impairment associated with FASD. A better understanding of these molecules and their effects on the developing hippocampus may lead to novel treatment strategies for FASD.

The wmN1 Enhancer Region of the Mouse Myelin Proteolipid Protein Gene (mPlp1) is Indispensable for Expression of an mPlp1-lacZ Transgene in Both the CNS and PNS.

The myelin proteolipid protein gene (PLP1) encodes the most abundant protein in CNS myelin. Expression of the gene must be strictly regulated, as evidenced by human X-linked leukodystrophies resulting from variations in PLP1 copy number, including elevated dosages as well as deletions. Recently, we showed that the wmN1 region in human PLP1 (hPLP1) intron 1 is required to promote high levels of an hPLP1-lacZ transgene in mice, using a Cre-lox approach. The current study tests whether loss of the wmN1 region from a related transgene containing mouse Plp1 (mPlp1) DNA produces similar results. In addition, we investigated the effects of loss of another region (ASE) in mPlp1 intron 1. Previous studies have shown that the ASE is required to promote high levels of mPlp1-lacZ expression by transfection analysis, but had no effect when removed from the native gene in mouse. Whether this is due to compensation by another regulatory element in mPlp1 that was not included in the mPlp1-lacZ constructs, or to differences in methodology, is unclear. Two transgenic mouse lines were generated that harbor mPLP(+)Z/FL. The parental transgene utilizes mPlp1 sequences (proximal 2.3kb of 5'-flanking DNA to the first 37bp of exon 2) to drive expression of a lacZ reporter cassette. Here we demonstrate that mPLP(+)Z/FL is expressed in oligodendrocytes, oligodendrocyte precursor cells, olfactory ensheathing cells and neurons in brain, and Schwann cells in sciatic nerve. Loss of the wmN1 region from the parental transgene abolished expression, whereas removal of the ASE had no effect.

The wmN1 enhancer region in intron 1 is required for expression of human PLP1.

The myelin proteolipid protein gene (PLP1) encodes the most abundant protein present in myelin from the central nervous system (CNS). Its expression must be tightly controlled as evidenced by mutations that alter PLP1 dosage; both overexpression (elevated PLP1 copy number) and lack thereof (PLP1 deletion) result in X-linked genetic disorders in man. However, not much is known about the mechanisms that govern expression of the human gene. To address this, transgenic mice were generated which utilize human PLP1 (hPLP1) sequences (proximal 6.2 kb of 5'-flanking DNA to the first 38 bp of exon 2) to drive expression of a lacZ reporter cassette. LoxP sites were incorporated around a 1.5-kb section of hPLP1 intron 1 since it contains sequence orthologous to the wmN1 region from mouse which, previously, was shown to augment expression of a minimally-promoted transgene coincident with the active myelination period of CNS development. Eight transgenic lines were generated with the parental, 6.2hPLP(+)Z/FL, transgene. All lines expressed the transgene appropriately in brain as evidenced by staining with X-gal in white matter regions and olfactory bulb. Removal of the "wmN1" region from 6.2hPLP(+)Z/FL with a ubiquitously expressed Cre-driver caused a dramatic reduction in transgene activity. These results demonstrate for the first time that the wmN1 enhancer region: (1) is functional in hPLP1; (2) works in collaboration with its native promoter-not just a basal heterologous promoter; (3) is required for high levels of hPLP1 gene activity; (4) has a broader effect, both spatially and temporally, than originally projected with mPlp1.

Effects of Intron 1 Sequences on Human PLP1 Expression: Implications for PLP1-Related Disorders.

Alterations in the myelin proteolipid protein gene (PLP1) may result in rare X-linked disorders in humans such as Pelizaeus–Merzbacher disease and spastic paraplegia type 2. PLP1 expression must be tightly regulated since null mutations, as well as elevated PLP1 copy number, both lead to disease. Previous studies with Plp1-lacZ transgenic mice have demonstrated that mouse Plp1 (mPlp1) intron 1 DNA (which accounts for slightly more than half of the gene) is required for the mPlp1 promoter to drive significant levels of reporter gene expression in brain. However not much is known about the mechanisms that control expression of the human PLP1 gene (hPLP1). Therefore this review will focus on sequences in hPLP1 intron 1 DNA deemed important for hPLP1 gene activity as well as a couple of “human-specific” supplementary exons within the first intron which are utilized to generate novel splice variants, and the potential role that these sequences may play in PLP1-linked disorders.

Control of human PLP1 expression through transcriptional regulatory elements and alternatively spliced exons in intron 1.

*These authors contributed equally to this work.Although the myelin proteolipid protein gene (PLP1) encodes the most abundant protein in central nervous system (CNS) myelin, not much is known about the mechanisms that govern expression of the human gene (hPLP1). Much more is known about the processes that regulate Plp1 gene expression in rodents. From studies with Plp1-lacZ transgenic mice, it was determined that the first intron of mouse Plp1 (mPlp1) is required to attain high levels of expression in brain, concurrent with the active myelination period. Other studies have suggested that within mPlp1 intron 1 (>8 kb) lie several regions with enhancer-like activity. To test whether these sequences (and possibly others) in hPLP1 intron 1 are functional, deletion-transfection analysis was performed with hPLP1-lacZ constructs that contain various portions of the intron, or lack it altogether. Results presented here demonstrate the importance of hPLP1 intron 1 in achieving maximal levels of expression in the immortalized oligodendroglial cell line, Oli-neu. Deletion analysis indicates that the intron contains multiple positive regulatory elements which are active in Oli-neu cells. Some of these elements appear to be functionally conserved between human and mouse, while others are not. Furthermore, our studies demonstrate that multiple splice variants can be formed due to inclusion of extra (supplementary) exons from what is classically thought of as hPLP1 intron 1. Thus, splicing of these novel exons (which are not recognized as such in mPlp1 due to lack of conserved splice sites) must utilize factors common to both human and mouse since Oli-neu cells are of mouse origin.

Two-photon imaging of remyelination of spinal cord axons by engrafted neural precursor cells in a viral model of multiple sclerosis.

Neural precursor cells (NPCs) offer a promising approach for treating demyelinating diseases. However, the cellular dynamics that underlie transplanted NPC-mediated remyelination have not been described. Using two-photon imaging of a newly developed ventral spinal cord preparation and a viral model of demyelination, we describe the motility and intercellular interactions of transplanted mouse NPCs expressing green fluorescent protein (GFP) with damaged axons expressing yellow fluorescent protein (YFP). Our findings reveal focal axonal degeneration that occurs in the ventral side of the spinal cord within 1 wk following intracranial instillation with the neurotropic JHM strain of mouse hepatitis virus (JHMV). Axonal damage precedes extensive demyelination and is characterized by swelling along the length of the axon, loss of YFP signal, and transected appearance. NPCs engrafted into spinal cords of JHMV-infected mice exhibited diminished migration velocities and increased proliferation compared with transplanted cells in noninfected mice. NPCs preferentially accumulated within areas of axonal damage, initiated direct contact with axons, and subsequently expressed the myelin proteolipid protein gene, initiating remyelination. These findings indicate that NPCs transplanted into an inflammatory demyelinating microenvironment participate directly in therapeutic outcome through the wrapping of myelin around damaged neurons.

Investigation of the proteolipid protein promoter activity during demyelination and repair

The transgenic plp-GFP mouse line expressing the green fluorescent protein (GFP) driven by the mouse myelin proteolipid protein (plp) gene promoter has been previously used to study the contribution of the plp lineage to oligodendrocyte development in the embryonic brain. Here, we show that the GFP fluorescence reflects the developmental expression of proteolipid protein during the postnatal development until adulthood in brain slices and in primary cultures of plp-GFP+ cells derived from postnatal animals. In the adult brain, plp-GFP-expressing cells are mature oligodendrocytes but not oligodendroglial progenitors. In the model of focal demyelination induced by lysolecithin (LPC) in the corpus callosum of adult plp-GFP animals, we observed an up-regulation of the morphogen Sonic Hedgehog (Shh) in the LPC-induced lesion but not in the control animals. Moreover, we show that the adenovirus-mediated transfer of Shh in the lesion results in the attenuation of the demyelination extent as evidenced by GFP fluorescence analysis in Shh-treated and control animals. Altogether these data show how plp-GFP fluorescence can be monitored to follow the oligodendrocyte lineage during demyelination and identify Shh morphogen as an important factor during repair.

Targeted deletion of the antisilencer/enhancer (ASE) element from intron 1 of the myelin proteolipid protein gene (Plp1) in mouse reveals that the element is dispensable for Plp1 expression in brain during development and remyelination

Myelin proteolipid protein gene (Plp1) expression is temporally regulated in brain, which peaks during the active myelination period of CNS development. Previous studies with Plp1-lacZ transgenic mice demonstrated that (mouse) Plp1 intron 1 DNA is required for high levels of expression in oligodendrocytes. Deletion-transfection analysis revealed the intron contains a single positive regulatory element operative in the N20.1 oligodendroglial cell line, which was named ASE (antisilencer/enhancer) based on its functional properties in these cells. To investigate the role of the ASE in vivo, the element was deleted from the native gene in mouse using a Cre/lox strategy. Although removal of the ASE from Plp1-lacZ constructs profoundly decreased expression in transfected oligodendroglial cell lines (N20.1 and Oli-neu), the element was dispensable to achieve normal levels of Plp1 gene expression in mouse during development (except perhaps at postnatal day15) and throughout the remyelination period following cuprizone-induced (acute) demyelination. Thus, it is possible that the ASE is non-functional in vivo, or that loss of the ASE from the native gene in mouse can be compensated for by the presence of other regulatory elements within the Plp1 gene.

Factor binding sites within the antisilencer/enhancer region of the myelin proteolipid protein gene are important for transcriptional activity in cultured oligodendroglial cells

The first intron of the myelin proteolipid protein (Plp) gene contains a positive regulatory region known as the antisilencer/enhancer (ASE) that is located between nucleotides 1083 and 1177 of the intron and is sufficient to restrict high‐level expression of the gene to oligodendrocytes in mice. To identify critical transcriptional control elements within the ASE, we have created a series of twelve non‐overlapping 8 bp linker scan mutations across the ASE region. These mutant ASE fragments were inserted in both orientations into a Plp‐lacZ fusion gene containing a shortened form of intron 1 that lacked the ASE. Relative transcriptional activities of these PLP‐lacZ constructs were assessed indirectly by measuring the β‐galactosidase activities in extracts of transiently transfected oligodendroglial (N20.1) cells. Five of the mutants with linkers inserted near the center of the ASE demonstrated reduced activity (10–50% of wild‐type levels). This effect was relatively independent of the orientation of the ASE inserted into the Plp‐lacZ constructs, consistent with the function of this region as a transcriptional enhancer. LS mutations at these same positions were previously shown to interfere with the binding of factors present in nuclear extracts of N20.1 cells, suggesting that these factors may be important in the oligodendrocyte‐specific regulation of the Plp gene. Supported by NIH grant P20 RR‐16460.

Mutation in the myelin proteolipid protein gene alters BK and SK channel function in the caudal medulla

Proteolipid protein (Plp) gene mutation in rodents causes severe CNS dysmyelination, early death, and lethal hypoxic ventilatory depression (Miller et al., 2004). To determine if Plp mutation alters neuronal function critical for control of breathing, the nucleus tractus solitarii (nTS) of four rodent strains were studied: myelin deficient rats (MD), myelin synthesis deficient (Plp(msd)), and Plp(null) mice, as well as shiverer (Mbp(shi)) mice, a myelin basic protein mutant. Current-voltage relationships were analyzed using whole-cell patch-clamp in 300 microm brainstem slices. Voltage steps were applied, and inward and outward currents quantified. MD, Plp(msd), and Plp(null), but not Mbp(shi) neurons exhibited reduced outward current in nTS at P21. Apamin blockade of SK calcium-dependent currents and iberiotoxin blockade of BK calcium-dependent currents in the P21 MD rat demonstrated reduced outward current due to dysfunction of these channels. These results provide evidence that Plp mutation specifically alters neuronal excitability through calcium-dependent potassium channels in nTS.

Mice with Altered Myelin Proteolipid Protein Gene Expression Display Cognitive Deficits Accompanied by Abnormal Neuron-Glia Interactions and Decreased Conduction Velocities

Conduction velocity (CV) of myelinated axons has been shown to be regulated by oligodendrocytes even after myelination has been completed. However, how myelinating oligodendrocytes regulate CV, and what the significance of this regulation is for normal brain function remain unknown. To address these questions, we analyzed a transgenic mouse line harboring extra copies of the myelin proteolipid protein 1 (plp1) gene (plp1(tg/-) mice) at 2 months of age. At this stage, the plp1(tg/-) mice have an unaffected myelin structure with a normally appearing ion channel distribution, but the CV in all axonal tracts tested in the CNS is greatly reduced. We also found decreased axonal diameters and slightly abnormal paranodal structures, both of which can be a cause for the reduced CV. Interestingly the plp1(tg/-) mice showed altered anxiety-like behaviors, reduced prepulse inhibitions, spatial learning deficits and working memory deficit, all of which are schizophrenia-related behaviors. Our results implicate that abnormalities in the neuron-glia interactions at the paranodal junctions can result in reduced CV in the CNS, which then induces behavioral abnormalities related to schizophrenia.

Different Proteolipid Protein Mutants Exhibit Unique Metabolic Defects

PMD (Pelizaeus–Merzbacher disease), a CNS (central nervous system) disease characterized by shortened lifespan and severe neural dysfunction, is caused by mutations of the PLP1 (X-linked myelin proteolipid protein) gene. The majority of human PLP1 mutations are caused by duplications; almost all others are caused by missense mutations. The cellular events leading to the phenotype are unknown. The same mutations in non-humans make them ideal models to study the mechanisms that cause neurological sequelae. In the present study we show that mice with Plp1 duplications (Plp1tg) have major mitochondrial deficits with a 50% reduction in ATP, a drastically reduced mitochondrial membrane potential and increased numbers of mitochondria. In contrast, the jp (jimpy) mouse with a Plp1 missense mutation exhibits normal mitochondrial function. We show that PLP in the Plp1tg mice and in Plp1-transfected cells is targeted to mitochondria. PLP has motifs permissive for insertion into mitochondria and deletions near its N-terminus prevent its co-localization to mitochondria. These novel data show that Plp1 missense mutations and duplications of the native Plp1 gene initiate uniquely different cellular responses.

Leydig cells express the myelin proteolipid protein gene and incorporate a new alternatively spliced exon

Although the myelin proteolipid protein gene ( Plp1) is highly expressed in the central nervous system encoding the most abundant myelin protein in oligodendrocytes, it is also expressed in other tissues, including testis. Transgenic studies with mice that harbor Plp1–lacZ fusion genes suggest that Leydig cells are the source of Plp1 gene expression in testis. However, virtually nothing is known about Plp1 gene regulation in Leydig cells, which is the focus of this study. The first intron contains both positive and negative regulatory elements that are important in regulating Plp1 gene expression in oligodendrocytes. To test whether these elements are functional in Leydig cells, a battery of Plp1–lacZ fusion genes with partial deletion of Plp1 intron 1 sequence was transfected into the mouse Leydig cell line, TM3. Results presented here suggest that an enhancer, which is very potent in oligodendrocytes, is only nominally active in TM3 cells. The intron also contains several negative regulatory elements that are operative in TM3 cells. Moreover a new exon (exon 1.2) was identified within the first ‘intron’ resulting in novel splice variants in TM3 cells. Western blot analysis suggests that these splice variants, along with those containing another alternatively spliced exon (exon 1.1) derived from intron 1 sequence, give rise to multiple Plp1 gene products in the mouse testis.

Localizing factor binding sites within an intronic enhancer of the myelin proteolipid protein gene

Localizing factor binding sites within an intronic enhancer of the myelin proteolipid protein gene

Proteomic analysis of nuclear factors binding to an intronic enhancer in the myelin proteolipid protein gene

The myelin proteolipid protein gene (Plp1) encodes the most abundant protein found in CNS myelin, accounting for nearly one-half of the total protein. Its expression in oligodendrocytes is developmentally regulated - peaking during the active myelination period of CNS development. Previously, we have identified a novel enhancer (designated ASE) in intron 1 DNA that appears to be important in mediating the surge of Plp1 gene activity during the active myelination period. Evidence suggests that the ASE participates in the formation of a specialized multi-protein/DNA complex called an enhanceosome. The current study describes an optimized, five-step, DNA affinity chromatography purification procedure to purify nuclear proteins from mouse brain that bind to the 85-bp ASE sequence, specifically. Electrophoretic mobility shift assay analysis demonstrated that specific DNA-binding activity was retained throughout the purification procedure, resulting in concomitant enrichment of nucleoprotein complexes. Identification of the purported regulatory factors was achieved through mass spectrometry analysis and included over 20 sequence-specific DNA-binding proteins. Supplementary western blot analyses to determine which of these sequence-specific factors are present in oligodendrocytes, and their developmental and regional expression in whole brain, suggest that Puralpha and Purbeta rank highest among the candidate factors as constituents of the multi-protein complex formed on the ASE.

Processing of PLP in a model of Pelizaeus‐Merzbacher disease/SPG2 due to the rumpshaker mutation

The rumpshaker mutation of the X-linked myelin proteolipid protein (PLP1) gene causes spastic paraplegia type 2 or a mild form of Pelizaeus-Merzbacher disease in man. The identical mutation occurs spontaneously in mice. Both human and murine diseases are associated with dysmyelination. Using the mouse model, we show that the low steady state levels of PLP result from accelerated proteasomal degradation rather than decreased synthesis. The T(1/2) for degradation of rumpshaker PLP is 11 h compared with 23 h for wild type. A minority of newly synthesized PLP is incorporated into myelin in the correct orientation but at a reduced rate compared with wild type. However, inhibition of proteasomal degradation does not increase the level of PLP incorporated into myelin. As Plp null mice do not have a similar myelin deficiency, it is unlikely that the reduced PLP levels are the main cause of the dysmyelination. Rumpshaker oligodendrocytes also have a reduced level of other myelin proteins, such as MBP, although the mechanisms are not yet defined but are likely to operate at a translational or post-translational level.

Characterization of an intronic enhancer that regulates myelin proteolipid protein (Plp) gene expression in oligodendrocytes

The myelin proteolipid protein (Plp) gene is expressed in oligodendrocytes and encodes the most abundant protein (approximately 50%) present in mature myelin from the central nervous system (CNS). Plp gene activity is low to nonexistent early in development but sharply increases, concurrently with the active myelination period of CNS development. Work from our laboratory suggests that the temporal regulation of Plp gene expression in mice is mediated by a positive regulatory element located within Plp intron 1 DNA. We have termed this regulatory element/region ASE (for antisilencer/enhancer). The ASE is situated approximately 1 kb downstream of exon 1 DNA and encompasses nearly 100 bp. To understand the mechanisms by which the ASE augments Plp gene expression in oligodendrocytes, Plp-lacZ constructs were generated and transfected into a mouse oligodendroglial cell line (N20.1). Results presented here demonstrate that upstream regulatory elements in the Plp promoter/5'-flanking DNA are not required for ASE activity; the ASE worked perfectly well when the thymidine kinase (TK) promoter was substituted for the Plp promoter. However, the relative location of the ASE appears to be important. When placed upstream of 2.4 kb of Plp 5'-flanking DNA, or downstream of the lacZ expression cassette, the ASE was no longer effective. Thus, the ASE might have to be in the context of the intron in order to function. To begin to identify the crucial nucleotides within the ASE, orthologous sequences from rat, human, cow, and pig Plp genes were swapped for the mouse sequence. Results presented here demonstrate that the orthologous sequence from rat can substitute for the mouse ASE, unlike those from human, cow, or pig.

The myelin proteolipid protein gene modulates apoptosis in neural and non-neural tissues.

Mutations of the myelin proteolipid protein gene (Plp) are associated with excessive programmed cell death (PCD) of oligodendrocytes. We show for the first time that PLP is a molecule ubiquitously expressed in non-neural tissues during normal development, and that the level of native PLP modulates the level of PCD. We analyze three non-neural tissues, and show that native PLP is expressed in trophoblasts, spermatogonia, and cells of interdigital webbing. The non-neural cells that express high levels of native PLP also undergo PCD. The level of PLP expression modulates the level of PCD because mice that overexpress native PLP have increased PCD and mice deficient in PLP have decreased PCD. We show that overexpression of native PLP causes a dramatic acidification of extracellular fluid that, in turn, causes increased PCD. These studies show that the level of native PLP modulates the amount of PCD during normal development via a pH-dependent mechanism.