Deletion Of LIS1 Research Articles

The Cytoplasmic Dynein Associated Protein NDE1 Regulates Osteoclastogenesis by Modulating M-CSF and RANKL Signaling Pathways.

Cytoskeleton organization and lysosome secretion play an essential role in osteoclastogenesis and bone resorption. The cytoplasmic dynein is a molecular motor complex that regulates microtubule dynamics and transportation of cargos/organelles, including lysosomes along the microtubules. LIS1, NDE1, and NDEL1 belong to an evolutionary conserved pathway that regulates dynein functions. Disruption of the cytoplasmic dynein complex and deletion of LIS1 in osteoclast precursors arrest osteoclastogenesis. Nonetheless, the role of NDE1 and NDEL1 in osteoclast biology remains elusive. In this study, we found that knocking-down Nde1 expression by lentiviral transduction of specific shRNAs markedly inhibited osteoclastogenesis in vitro by attenuating the proliferation, survival, and differentiation of osteoclast precursor cells via suppression of signaling pathways downstream of M-CSF and RANKL as well as osteoclast differentiation transcription factor NFATc1. To dissect how NDEL1 regulates osteoclasts and bone homeostasis, we generated Ndel1 conditional knockout mice in myeloid osteoclast precursors (Ndel1ΔlysM) by crossing Ndel1-floxed mice with LysM-Cre mice on C57BL/6J background. The Ndel1ΔlysM mice developed normally. The µCT analysis of distal femurs and in vitro osteoclast differentiation and functional assays in cultures unveiled the similar bone mass in both trabecular and cortical bone compartments as well as intact osteoclastogenesis, cytoskeleton organization, and bone resorption in Ndel1ΔlysM mice and cultures. Therefore, our results reveal a novel role of NDE1 in regulation of osteoclastogenesis and demonstrate that NDEL1 is dispensable for osteoclast differentiation and function.

Hepatic loss of Lissencephaly 1 (Lis1) induces fatty liver and accelerates liver tumorigenesis in mice

The liver is a major organ in lipid metabolism, and its malfunction leads to various diseases. Nonalcoholic fatty liver disease, the most common chronic liver disorder in developed countries, is characterized by the abnormal retention of excess lipid within hepatocytes and predisposes individuals to liver cancer. We previously reported that the levels of Lissencephaly 1 (LIS1, also known as PAFAH1B1) are down-regulated in human hepatocellular carcinoma. Following up on this observation, we found that genetic deletion of Lis1 in the mouse liver increases lipid accumulation and inflammation in this organ. Further analysis revealed that loss of Lis1 triggers endoplasmic reticulum (ER) stress and reduces triglyceride secretion. Attenuation of ER stress by addition of tauroursodeoxycholic acid (TUDCA) diminished lipid accumulation in the Lis1-deficient hepatocytes. Moreover, the Golgi stacks were disorganized in Lis1-deficient liver cells. Of note, the Lis1 liver-knockout mice exhibited increased hepatocyte ploidy and accelerated development of liver cancer after exposure to the liver carcinogen diethylnitrosamine (DEN). Taken together, these findings suggest that reduced Lis1 levels can spur the development of liver diseases from steatosis to liver cancer and provide a useful model for delineating the molecular pathways that lead to these diseases.

The LIS1/NDE1 Complex Is Essential for FGF Signaling by Regulating FGF Receptor Intracellular Trafficking

Intracellular transport of membranous organelles and protein complexes to various destinations is fundamental to signaling transduction and cellular function. The cytoplasmic dynein motor and its regulatory proteins LIS1 and NDE1 are required for transporting a variety of cellular cargos along the microtubule network. In this study, we show that deletion of Lis1 in developing lung endoderm and limb mesenchymal cells causes agenesis of the lungs and limbs. In bothmutants, there is increased cell death and decreased fibroblast growth factor (FGF) signaling activity. Mechanistically, LIS1 and its interacting protein NDE1/NDEL1 are associated with FGF receptor-containing vesicles and regulate FGF receptor intracellular trafficking and degradation. Notably, FGF signaling promotes NDE1 tyrosine phosphorylation, which leads to dissociation of LIS1/NDE1 complex. Thus, our studies identify the LIS1/NDE1 complex as an important FGF signaling regulator and provide insights into the bidirectional regulation of cell signaling and transport machinery for endocytosis.

The Microtubule-Associated Protein Lis1 Regulates T Lymphocyte Homeostasis and Differentiation.

The microtubule-associated protein lissencephaly 1 (Lis1) is a key regulator of cell division during stem cell renewal and differentiation. In this study, we examined the role of Lis1 in T lymphocyte homeostasis and fate diversification in response to microbial infection. T cell-specific deletion of Lis1 resulted in depletion of the peripheral CD4(+) and CD8(+) T lymphocyte pool owing to a loss of homeostatic, cytokine-induced proliferation. In contrast, cognate Ag-triggered proliferation was much less affected, enabling Lis1-deficient CD8(+) T cells to differentiate into terminal effector cells in response to microbial infection. Strikingly, however, the specification of Lis1-deficient long-lived memory CD8(+) T lymphocytes was impaired due, in part, to an apparent failure to differentiate appropriately to IL-15. Taken together, these findings suggest that Lis1 plays an important role in T cell homeostasis and the generation of memory T lymphocytes.

The microtubule-associated protein Lis1 regulates T lymphocyte homeostasis and differentiation

Abstract The microtubule-associated protein Lissencephaly 1 (Lis1) is a key regulator of cell division during stem cell renewal and differentiation. We examined the role of Lis1 in T lymphocyte homeostasis and fate diversification in response to microbial infection. T cell-specific deletion of Lis1 resulted in depletion of the peripheral CD4+ and CD8+ T lymphocyte pool, owing to a selective loss of homeostatic, cytokine-induced proliferation. By contrast, cognate antigen-triggered proliferation was relatively unaffected in CD8+ T cells, enabling Lis1-deficient T cells to differentiate into terminal effector cells in response to microbial infection. Strikingly, however, Lis1-deficient CD8+ T cells failed to develop into long-lived memory lymphocytes. Taken together, these findings suggest that Lis1 plays an important role in T cell homeostasis and the generation of memory T lymphocytes.

LIS1 Regulates Osteoclastogenesis through Modulation of M-SCF and RANKL Signaling Pathways and CDC42.

We have previously reported that depletion of LIS1, a key regulator of microtubules and cytoplasmic dynein motor complex, in osteoclast precursor cells by shRNAs attenuates osteoclastogenesis in vitro. However, the underlying mechanisms remain unclear. In this study, we show that conditional deletion of LIS1 in osteoclast progenitors in mice led to increased bone mass and decreased osteoclast number on trabecular bone. In vitro mechanistic studies revealed that loss of LIS1 had little effects on cell cycle progression but accelerated apoptosis of osteoclast precursor cells. Furthermore, deletion of LIS1 prevented prolonged activation of ERK by M-CSF and aberrantly enhanced prolonged JNK activation stimulated by RANKL. Finally, lack of LIS1 abrogated M-CSF and RANKL induced CDC42 activation and retroviral transduction of a constitutively active form of CDC42 partially rescued osteoclastogenesis in LIS1-deficient macrophages. Therefore, these data identify a key role of LIS1 in regulation of cell survival of osteoclast progenitors by modulating M-CSF and RANKL induced signaling pathways and CDC42 activation.

Lis1 regulates asymmetric division in hematopoietic stem cells and in leukemia.

Cell fate can be controlled through asymmetric division and segregation of protein determinants. But the regulation of this process in the hematopoietic system is poorly understood. Here we show that the dynein binding protein Lis1 (Pafah1b1) is critically required for blood formation and hematopoietic stem cell function. Conditional deletion of Lis1 in the hematopoietic system led to a severe bloodless phenotype, depletion of the stem cell pool and embryonic lethality. Further, the loss of Lis1 accelerated cell differentiation, in part through defects in spindle positioning and inheritance of cell fate determinants. Finally, deletion of Lis1 blocked propagation of myeloid leukemia and led to a marked improvement in animal survival, suggesting that Lis1 is also required for oncogenic growth. These data identify a key role for Lis1 in hematopoietic stem cells, and mark the directed control of asymmetric division as a critical regulator of normal and malignant hematopoietic development.

Lis1 mediates planar polarity of auditory hair cells through regulation of microtubule organization

The V-shaped hair bundles atop auditory hair cells and their uniform orientation are manifestations of epithelial planar cell polarity (PCP) required for proper perception of sound. PCP is regulated at the tissue level by a conserved core Wnt/PCP pathway. However, the hair cell-intrinsic polarity machinery is poorly understood. Recent findings implicate hair cell microtubules in planar polarization of hair cells. To elucidate the microtubule-mediated polarity pathway, we analyzed Lis1 function in the auditory sensory epithelium in the mouse. We show that conditional deletion of Lis1 in developing hair cells causes defects in cytoplasmic dynein and microtubule organization, resulting in planar polarity defects without overt effects on the core PCP pathway. Lis1 ablation during embryonic development results in defects in hair bundle morphology and orientation, cellular organization and junctional nectin localization. We present evidence that Lis1 regulates localized Rac-PAK signaling in embryonic hair cells, probably through microtubule-associated Tiam1, a guanine nucleotide exchange factor for Rac. Lis1 ablation in postnatal hair cells significantly disrupts centrosome anchoring and the normal V-shape of hair bundles, accompanied by defects in the pericentriolar matrix and microtubule organization. Lis1 is also required for proper positioning of the Golgi complex and mitochondria as well as for hair cell survival. Together, our results demonstrate that Lis1 mediates the planar polarity of hair cells through regulation of microtubule organization downstream of the tissue polarity pathway.

Lis1 mediates planar polarity of auditory hair cells through regulation of microtubule organization

The V-shaped hair bundles atop auditory hair cells and their uniform orientation are manifestations of epithelial planar cell polarity (PCP) required for proper perception of sound. PCP is regulated at the tissue level by a conserved core Wnt/PCP pathway. However, the hair cell-intrinsic polarity machinery is poorly understood. Recent findings implicate hair cell microtubules in planar polarization of hair cells. To elucidate the microtubule-mediated polarity pathway, we analyzed Lis1 function in the auditory sensory epithelium in the mouse. We show that conditional deletion of Lis1 in developing hair cells causes defects in cytoplasmic dynein and microtubule organization, resulting in planar polarity defects without overt effects on the core PCP pathway. Lis1 ablation during embryonic development results in defects in hair bundle morphology and orientation, cellular organization and junctional nectin localization. We present evidence that Lis1 regulates localized Rac-PAK signaling in embryonic hair cells, probably through microtubule-associated Tiam1, a guanine nucleotide exchange factor for Rac. Lis1 ablation in postnatal hair cells significantly disrupts centrosome anchoring and the normal V-shape of hair bundles, accompanied by defects in the pericentriolar matrix and microtubule organization. Lis1 is also required for proper positioning of the Golgi complex and mitochondria as well as for hair cell survival. Together, our results demonstrate that Lis1 mediates the planar polarity of hair cells through regulation of microtubule organization downstream of the tissue polarity pathway.

LIS1 Deficiency Promotes Dysfunctional Synaptic Integration of Granule Cells Generated in the Developing and Adult Dentate Gyrus

Type I lissencephaly, a neuronal migration disorder characterized by cognitive disability and refractory epilepsy, is often caused by heterozygous mutations in the LIS1 gene. Histopathologies of malformation-associated epilepsies have been well described, but it remains unclear whether hyperexcitability is attributable to disruptions in neuronal organization or abnormal circuit function. Here, we examined the effect of LIS1 deficiency on excitatory synaptic function in the dentate gyrus of hippocampus, a region believed to serve critical roles in seizure generation and learning and memory. Mice with heterozygous deletion of LIS1 exhibited robust granule cell layer dispersion, and adult-born granule cells labeled with enhanced green fluorescent protein were abnormally positioned in the molecular layer, hilus, and granule cell layer. In whole-cell patch-clamp recordings, reduced LIS1 function was associated with greater excitatory synaptic input to mature granule cells that was consistent with enhanced release probability at glutamatergic synapses. Adult-born granule cells that were ectopically positioned in the molecular layer displayed a more rapid functional maturation and integration into the synaptic network compared with newborn granule cells located in the hilus or granule cell layer or in wild-type controls. In a conditional knock-out mouse, induced LIS1 deficiency in adulthood also enhanced the excitatory input to granule cells in the absence of neuronal disorganization. These findings indicate that disruption of LIS1 has direct effects on excitatory synaptic transmission independent of laminar disorganization, and the ectopic position of adult-born granule cells within a malformed dentate gyrus critically influences their functional maturation and integration.

Expression and function on embryonic development of lissencephaly-1 genes in zebrafish

Lissencephaly is a severe disease characterized by brain malformation. The main causative gene of lissencephaly is LIS1. Mutation or deletion of LIS1 leads to proliferation and migration deficiency of neurons in brain development. However, little is known about its biological function in embryonic development. In this article, we identified the expression patterns of zebrafish LIS1 gene and investigated its function in embryonic development. We demonstrated that zebrafish consisted of two LIS1 genes, LIS1a and LIS1b. Bioinformatics analysis revealed that LIS1 genes were conserved in evolution both in protein sequences and genomic structures. The expression patterns of zebrafish LIS1a and LIS1b showed that both transcripts were ubiquitously expressed at all embryonic developmental stages and in adult tissues examined. At the protein level, the LIS1 products mainly exist in brain tissue and in embryos at early stages as shown by western blotting analysis. The whole-mount immunostaining data showed that LIS1 proteins were distributed all over the embryos from 1-cell stage to 5 day post-fertilization. Knockdown of LIS1 protein expression through morpholino antisense oligonucleotides resulted in many developmental deficiencies in zebrafish, including brain malformation, circulation abnormality, and body curl. Taken together, our study suggested that zebrafish LIS1 plays a very important role in embryonic development.

Aberrant dentate gyrus cytoarchitecture and fiber lamination in LIS1 mutant mice

Mutant mice with a heterozygous deletion of LIS1, show varying degrees of hippocampal abnormality and enhanced excitability. To examine how LIS1 affects cytoarchitecture and fiber lamination in dentate gyrus (DG), we performed a series of immunohistochemistry studies. By using different neuronal- and glial-specific antibodies, we found that the majority of hippocampal cell populations were affected by heterozygous mutation of LIS1; some reelin-positive Cajal-Retzius cells were left undisturbed. Granule cell dispersion was significant in hippocampal sections from Lis1-deficient mice. However, the fiber termination of commissural/associational fibers and mossy fibers appeared relatively compact despite obvious granule cell dispersion and CA1-CA3 pyramidal cell disorganization. vGlut1-immunoreactive axon terminals were found aberrantly traversing the dispersed granule cell layer. Consistent with previous observations, we also found that immature granule cells in Lis1 mutants, here stained with antibodies to doublecortin (DCX) and Mash-1, are aberrantly located and bear an abnormal cellular morphology. Our findings suggest that LIS1 mutants exhibit abnormal cell positioning and aberrant hippocampal neurogenesis, but maintain relatively normal fiber termination patterns. The functional consequences of hippocampal granule cell dispersion could offer critical insight to the epileptic and cognitive disorder associated with LIS1 haploinsufficiency.

Neuroepithelial Stem Cell Proliferation Requires LIS1 for Precise Spindle Orientation and Symmetric Division

Mitotic spindle orientation and plane of cleavage in mammals is a determinant of whether division yields progenitor expansion and/or birth of new neurons during radial glial progenitor cell (RGPC) neurogenesis, but its role earlier in neuroepithelial stem cells is poorly understood. Here we report that Lis1 is essential for precise control of mitotic spindle orientation in both neuroepithelial stem cells and radial glial progenitor cells. Controlled gene deletion of Lis1 in vivo in neuroepithelial stem cells, where cleavage is uniformly vertical and symmetrical, provokes rapid apoptosis of those cells, while radial glial progenitors are less affected. Impaired cortical microtubule capture via loss of cortical dynein causes astral and cortical microtubules to be greatly reduced in Lis1-deficient cells. Increased expression of the LIS/dynein binding partner NDEL1 restores cortical microtubule and dynein localization in Lis1-deficient cells. Thus, control of symmetric division, essential for neuroepithelial stem cell proliferation, is mediated through spindle orientation determined via LIS1/NDEL1/dynein-mediated cortical microtubule capture.

Partial Deletion of LIS1: A Pitfall in Molecular Diagnosis of Miller-Dieker Syndrome

Miller-Dieker syndrome represents a microdeletion syndrome spanning the LIS1 locus at 17p13.3, the deletion of which leads to lissencephaly. A fluorescence in situ hybridization study using an LIS1 probe is considered the standard laboratory diagnostic method for Miller-Dieker syndrome. This report documents a Miller-Dieker syndrome patient who tested normal when a commercially available LIS1 fluorescence in situ hybridization study probe was used but was later demonstrated to have a partial deletion of the LIS1 locus. The present case exemplifies a major shortcoming of commercially available fluorescence in situ hybridization studies for the diagnosis of microdeletion syndromes such as Miller-Dieker syndrome: that is, relatively small deletion can potentially remain undetected.

Granule Cell Dispersion and Aberrant Neurogenesis in the Adult Hippocampus of an LIS1 Mutant Mouse

Human type 1 lissencephaly is a severe brain malformation associated with cognitive dysfunction and intractable epilepsy. Mutant mice with a heterozygous deletion of LIS1 show varying degrees of hippocampal abnormality and enhanced excitability. Whether a reduction of LIS1 function affects adult hippocampal neurogenesis, and if so, whether aberrant neurogenesis contributes to the generation of a disorganized hippocampus remain unknown. Previous reports indicate the presence of multiple pyramidal cell layers and granule cell dispersion in LIS1 mutant mice. Here we observed disruption of the subgranular zone and glial fibrillary acidic protein-immunoreactive radial astrocytes in the dentate gyrus of adult LIS1 mice. Using pulse-chase bromodeoxyuridine (BrdU) labeling combined with neuronal and glial antibody staining we provide evidence for ectopic adult neurogenesis in LIS1 mice. A gradually decreased survival rate for these newborn granule cells was also demonstrated in LIS1 mice 7 days after BrdU injection. This reduced survival rate was associated with impaired neuronal differentiation 28 days after BrdU administration. Thus, LIS1 haploinsufficiency can lead to abnormal cell proliferation, migration and differentiation in the adult dentate gyrus.

Refinement of a 400-kb Critical Region Allows Genotypic Differentiation between Isolated Lissencephaly, Miller-Dieker Syndrome, and Other Phenotypes Secondary to Deletions of 17p13.3

Deletions of 17p13.3, including the LIS1 gene, result in the brain malformation lissencephaly, which is characterized by reduced gyration and cortical thickening; however, the phenotype can vary from isolated lissencephaly sequence (ILS) to Miller-Dieker syndrome (MDS). At the clinical level, these two phenotypes can be differentiated by the presence of significant dysmorphic facial features and a more severe grade of lissencephaly in MDS. Previous work has suggested that children with MDS have a larger deletion than those with ILS, but the precise boundaries of the MDS critical region and causative genes other than LIS1 have never been fully determined. We have completed a physical and transcriptional map of the 17p13.3 region from LIS1 to the telomere. Using fluorescence in situ hybridization, we have mapped the deletion size in 19 children with ILS, 11 children with MDS, and 4 children with 17p13.3 deletions not involving LIS1. We show that the critical region that differentiates ILS from MDS at the molecular level can be reduced to 400 kb. Using somatic cell hybrids from selected patients, we have identified eight genes that are consistently deleted in patients classified as having MDS. In addition, deletion of the genes CRK and 14-3-3 epsilon delineates patients with the most severe lissencephaly grade. On the basis of recent functional data and the creation of a mouse model suggesting a role for 14-3-3 epsilon in cortical development, we suggest that deletion of one or both of these genes in combination with deletion of LIS1 may contribute to the more severe form of lissencephaly seen only in patients with MDS.

Hippocampal Abnormalities and Enhanced Excitability in a Murine Model of Human Lissencephaly

Human cortical heterotopia and neuronal migration disorders result in epilepsy; however, the precise mechanisms remain elusive. Here we demonstrate severe neuronal dysplasia and heterotopia throughout the granule cell and pyramidal cell layers of mice containing a heterozygous deletion of Lis1, a mouse model of human 17p13.3-linked lissencephaly. Birth-dating analysis using bromodeoxyuridine revealed that neurons in Lis1+/- murine hippocampus are born at the appropriate time but fail in migration to form a defined cell layer. Heterotopic pyramidal neurons in Lis1+/- mice were stunted and possessed fewer dendritic branches, whereas dentate granule cells were hypertrophic and formed spiny basilar dendrites from which the principal axon emerged. Both somatostatin- and parvalbumin-containing inhibitory neurons were heterotopic and displaced into both stratum radiatum and stratum lacunosum-moleculare. Mechanisms of synaptic transmission were severely disrupted, revealing hyperexcitability at Schaffer collateral-CA1 synapses and depression of mossy fiber-CA3 transmission. In addition, the dynamic range of frequency-dependent facilitation of Lis1+/- mossy fiber transmission was less than that of wild type. Consequently, Lis1+/- hippocampi are prone to interictal electrographic seizure activity in an elevated [K(+)](o) model of epilepsy. In Lis1+/- hippocampus, intense interictal bursting was observed on elevation of extracellular potassium to 6.5 mM, a condition that resulted in only minimal bursting in wild type. These anatomical and physiological hippocampal defects may provide a neuronal basis for seizures associated with lissencephaly.

LIS1 and XLIS (DCX) mutations cause most classical lissencephaly, but different patterns of malformation.

Classical lissencephaly (LIS) is a neuronal migration disorder resulting in brain malformation, epilepsy and mental retardation. Deletions or mutations of LIS1 on 17p13.3 and mutations in XLIS ( DCX ) on Xq22.3-q23 produce LIS. Direct DNA sequencing of LIS1 and XLIS was performed in 25 children with sporadic LIS and no deletion of LIS1 by fluorescence in situ hybridization. Mutations of LIS1 were found by sequencing ( n = 8) and Southern blot ( n = 2) in a total of 10 patients (40%) of both sexes and mutations of XLIS in five males (20%). Combined with previous data, deletions or mutations of these two genes account for approximately 76% of isolated LIS. These data demonstrate that LIS1 and XLIS mutations cause the majority of, though not all, human LIS. The mutations in LIS1 were predicted to result in protein truncation in six of eight patients and splice site mutations in two, all of which disrupt one or more of the seven WD40 repeats contained in the LIS1 protein. Point mutations in XLIS identified the C-terminal serine/proline-rich region as potentially important for protein function. The patients with mutations were included in a genotype-phenotype analysis of 32 subjects with deletions or other mutations of these two genes. Whereas the brain malformation due to LIS1 mutations was more severe over the parietal and occipital regions, XLIS mutations produced the reverse gradient, which was more severe over the frontal cortex. The distinct LIS patterns suggest that LIS1 and XLIS may be part of overlapping, but distinct, signaling pathways that promote neuronal migration.

Platelet-Activating Factor Acetylhydrolase Expression and Activity Suggest a Link between Neuronal Migration and Platelet-Activating Factor

A hemizygous deletion ofLIS1,the gene encoding αLis1protein, causes Miller–Dieker syndrome (MDS). MDS is a developmental disorder characterized by neuronal migration defects resulting in a disorganization of the cerebral and cerebellar cortices. αLis1binds to two other proteins (β and γ) to form a heterotrimeric cytosolic enzyme which hydrolyzes platelet-activating factor (PAF). The existence of heterotrimers is implicated from copurification and crosslinking studies carried outin vitro.To determine whether such a heterotrimeric complex could be present in tissues, we have investigated whether the αLis1, β, and γ genes are coexpressed in the developing and adult brain. We have isolated murine cDNAs and show byin situhybridization that in developing brain tissues αLis1, β, and γ genes are coexpressed. This suggests that αLis1, β, and γ gene products form heterotrimers in developing neuronal tissues. In the adult brain, αLis1and β mRNAs continue to be coexpressed at high levels while γ gene expression is greatly diminished. This reduction in γ transcript levels is likely to result in a decline of the cellular concentration of αLis1, β, and γ heterotrimers. The developmental expression pattern of αLis1, β, and γ genes is consistent with the neuronal migration defects seen in MDS; regions containing migrating neurons such as the developing cerebral and cerebellar cortices express these genes at a particularly high level. Furthermore, we uncovered a correlation between γ gene expression, granule cell migration, and PAF hydrolytic activity in the cerebellum. In this tissue γ gene expression and PAF hydrolysis peaked at Postnatal Days P5 and P15, a period during which neuronal migration in the cerebellum is most extensive. Mechanisms by which PAF could affect neuronal migration are discussed.

Mutations in LIS1 ( ERG6) gene confer increased sodium and lithium uptake in Saccharomyces cerevisiae

A Saccharomyces cerevisiae mutant, lisI-1, hypersensitive to Li + and Na + was isolated from a wild-type strain after ethylmethane sulfonate mutagenesis. The rates of Li + and Na + uptake of the mutant are about 3–4-times higher than that of the wild-type; while the rates of cation efflux from the mutant and wild-type strains are indistinguishable. The LIS1 was isolated from a yeast genomic library by complementation of the cation hypersensitivity of the lisI-1 strain. LIS1 is a single copy, nonessential gene. However, the deletion of LIS1 from the wild-type results in a growth defect in addition to the cation hypersensitive phenotype. The order of increasing cation uptake rates of the wild-type and mutant strains, Δ 1::LEU2, correlates perfectly with the degree of cation hypersensitivity, suggesting that the cation hypersensitivity is primarily due to increased rates of cation influx. LIS1 encodes a membrane associated protein 384 amino acids long. Data base searches indicate that LIS1 is identical to ERG6 in S. cerevisiae which encodes a putative S-adenosylmethionine-dependent methyltransferase in the ergosterol biosynthetic pathway. Cell membranes of lis1 ( erg6) mutants are known to be devoid of ergosterol and have altered sterol composition. Since membrane sterols can influence the activity of cation transporters, the increased cation uptake of the lis1 mutants may stem from an altered function of one or many different membrane transporters.