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S Hekimi

Publications and source records attributed to S Hekimi.

At least 19 recordsLinked to original sources
PubMed2001

Ubiquinone is necessary for mouse embryonic development but is not essential for mitochondrial respiration.

Ubiquinone (UQ) is a lipid found in most biological membranes and is a co-factor in many redox processes including the mitochondrial respiratory chain. UQ has been implicated in protection from oxidative stress and in the aging process. Consequently, it is used as a dietary supplement and to treat mitochondrial diseases. Mutants of the clk-1 gene of the nematode Caenorhabditis elegans are fertile and have an increased life span, although they do not produce UQ but instead accumulate a biosynthetic intermediate, demethoxyubiquinone (DMQ). DMQ appears capable to partially replace UQ for respiration in vivo and in vitro. We have produced a vertebrate model of cells and tissues devoid of UQ by generating a knockout mutation of the murine orthologue of clk-1 (mclk1). We find that mclk1-/- embryonic stem cells and embryos accumulate DMQ instead of UQ. As in the nematode mutant, the activity of the mitochondrial respiratory chain of -/- embryonic stem cells is only mildly affected (65% of wild-type oxygen consumption). However, mclk1-/- embryos arrest development at midgestation, although earlier developmental stages appear normal. These findings indicate that UQ is necessary for vertebrate embryonic development but suggest that mitochondrial respiration is not the function for which UQ is essential when DMQ is present.

Animals↗
PubMed2001

Why only time will tell.

The nematode Caenorhabditis elegans has become a model system for the study of the genetic basis of aging. In particular, many mutations that extend life span have been identified in this organism. When loss-of-function mutations in a gene lead to life span extension, it is a necessary conclusion that the gene normally limits life span in the wild type. The effect of a given mutation depends on a number of environmental and genetic conditions. For example, the combination of two mutations can result in additive, synergistic, subtractive, or epistatic effects on life span. Valuable insight into the processes that determine life span can be obtained from such genetic analyses, especially when interpreted with caution, and when molecular information about the interacting genes is available. Thus, genetic and molecular analyses have implicated several genes classes (daf, clk and eat) in life span determination and have indicated that aging is affected by alteration of several biological processes, namely dormancy, physiological rates, food intake, and reproduction.

Aging↗
PubMed2001

Mouse CLK-1 is imported into mitochondria by an unusual process that requires a leader sequence but no membrane potential.

clk-1 has been identified and characterized in the nematode Caenorhabditis elegans as a gene that affects the rates, regularity, and synchrony of physiological processes. The CLK-1 protein is mitochondrial and is required for ubiquinone biosynthesis in yeast and in worms, but its biochemical function remains unclear. We have studied the expression of murine mclk1 in a variety of tissues, and we find that the pattern of mclk1 mRNA accumulation closely resembles that of mitochondrial genes involved in oxidative phosphorylation. The pattern of protein accumulation, however, is sharply distinct in some tissues; mCLK1 appears relatively enriched in the gut and depleted in the nervous tissue. We also show that mCLK1 is synthesized as a preprotein that is imported into the mitochondrial matrix, where a leader sequence is cleaved off and the protein becomes loosely associated with the inner membrane. However, in contrast to all known mitochondrial proteins that contain a cleavable pre-sequence, the import of mCLK1 does not require a mitochondrial membrane potential.

Amino Acid Sequence↗
PubMed2001

Altered quinone biosynthesis in the long-lived clk-1 mutants of Caenorhabditis elegans.

Mutations in the clk-1 gene of Caenorhabditis elegans result in an extended life span and an average slowing down of developmental and behavioral rates. However, it has not been possible to identify biochemical changes that might underlie the extension of life span observed in clk-1 mutants, and therefore the function of CLK-1 in C. elegans remains unknown. In this report, we analyzed the effect of clk-1 mutation on ubiquinone (UQ(9)) biosynthesis and show that clk-1 mutants mitochondria do not contain detectable levels of UQ(9). Instead, the UQ(9) biosynthesis intermediate, demethoxyubiquinone (DMQ(9)), is present at high levels. This result demonstrates that CLK-1 is absolutely required for the biosynthesis of UQ(9) in C. elegans. Interestingly, the activity levels of NADH-cytochrome c reductase and succinate-cytochrome c reductase in mutant mitochondria are very similar to those in the wild-type, suggesting that DMQ(9) can function as an electron carrier in the respiratory chain. To test this possibility, the short side chain derivative DMQ(2) was chemically synthesized. We find that DMQ(2) can act as an electron acceptor for both complex I and complex II in clk-1 mutant mitochondria, while another ubiquinone biosynthesis precursor, 3-hydroxy-UQ(2), cannot. The accumulation of DMQ(9) and its use in mutant mitochondria indicate, for the first time in any organism, a link between the alteration in the quinone species used in respiration and life span.

Animals↗
PubMed2001

Genetics of lifespan in C. elegans: molecular diversity, physiological complexity, mechanistic simplicity.

The nematode Caenorhabditis elegans is used as a model system for the study of aging. Several mutant strains that have an increased lifespan have been isolated and characterized genetically and molecularly. Molecular analysis reveals that diverse types of gene products can affect worm lifespan, including proteins active in signal transduction, transcription and silencing factors, mitochondrial enzymes, and at least one protein that affects telomere length. Genetic analysis, however, suggests that these activities all converge on a few key mechanisms that impinge on lifespan, namely the production, repair and prevention of molecular damage.

Animals↗
PubMed2001

Mitochondrial electron transport is a key determinant of life span in Caenorhabditis elegans.

Increased protection from reactive oxygen species (ROS) is believed to increase life span. However, it has not been clearly demonstrated that endogenous ROS production actually limits normal life span. We have identified a mutation in the Caenorhabditis elegans iron sulfur protein (isp-1) of mitochondrial complex III, which results in low oxygen consumption, decreased sensitivity to ROS, and increased life span. Furthermore, combining isp-1(qm150) with a mutation (daf-2) that increases resistance to ROS does not result in any significant further increase in adult life span. These findings indicate that both isp-1 and daf-2 mutations increase life span by lowering oxidative stress and result in the maximum life span increase that can be produced in this way.

Amino Acid Sequence↗
PubMed2001

Regulation of physiological rates in Caenorhabditis elegans by a tRNA-modifying enzyme in the mitochondria.

We show that the phenotype associated with gro-1(e2400) comprises the whole suite of features that characterize the phenotype of the clk mutants in Caenorhabditis elegans, including deregulated developmental, behavioral, and reproductive rates, as well as increased life span and a maternal effect. We cloned gro-1 and found that it encodes a highly conserved cellular enzyme, isopentenylpyrophosphate:tRNA transferase (IPT), which modifies a subset of tRNAs. In yeast, two forms of the enzyme are produced by alternative translation initiation, one of which is mitochondrial. In the gro-1 transcript there are also two possible initiator ATGs, between which there is a sequence predicted to encode a mitochondrial localization signal. A functional GRO-1::GFP fusion protein is localized diffusely throughout the cytoplasm and nucleus. A GRO-1::GFP initiated from the first methionine is localized exclusively to the mitochondria and rescues the mutant phenotype. In contrast, a protein initiated from the second methionine is localized diffusely throughout the cell and does not rescue the mutant phenotype. As oxygen consumption and ATP concentration have been reported to be unaffected in gro-1 mutants, our observations suggest that GRO-1 acts in mitochondria and regulates global physiology by unknown mechanisms.

Adenosine Triphosphate↗
PubMed2001

Phenotypic and suppressor analysis of defecation in clk-1 mutants reveals that reaction to changes in temperature is an active process in Caenorhabditis elegans.

Mutations in the Caenorhabditis elegans maternal-effect gene clk-1 affect cellular, developmental, and behavioral timing. They result in a slowing of the cell cycle, embryonic and postembryonic development, reproduction, and aging, as well as of the defecation, swimming, and pharyngeal pumping cycles. Here, we analyze the defecation behavior in clk-1 mutants, phenotypically and genetically. When wild-type worms are grown at 20 degrees and shifted to a new temperature, the defecation cycle length is significantly affected by that new temperature. In contrast, we find that when clk-1 mutants are shifted, the defecation cycle length is unaffected by that new temperature. We carried out a screen for mutations that suppress the slow defecation phenotype at 20 degrees and identified two distinct classes of genes, which we call dsc for defecation suppressor of clk-1. Mutations in one class also restore the ability to react normally to changes in temperature, while mutations in the other class do not. Together, these results suggest that clk-1 is necessary for readjusting the defecation cycle length in response to changes in temperature. On the other hand, in the absence of clk-1 activity, we observe temperature compensation, a mechanism that maintains a constant defecation period in the face of changes in temperature.

Animals↗
PubMed2001

The C. elegans maternal-effect gene clk-2 is essential for embryonic development, encodes a protein homologous to yeast Tel2p and affects telomere length.

The Caenorhabditis elegans maternal-effect clk genes are involved in the temporal control of development and behavior. We report the genetic and molecular characterization of clk-2. A temperature-sensitive mutation in the gene clk-2 affects embryonic and post-embryonic development, reproduction, and rhythmic behaviors. Yet, virtually all phenotypes are fully maternally rescued. Embryonic development strictly requires the activity of maternal clk-2 during a narrow time window between oocyte maturation and the two- to four-cell embryonic stage. Positional cloning of clk-2 reveals that it encodes a protein homologous to S. cerevisiae Tel2p. In yeast, the gene TEL2 regulates telomere length and participates in gene silencing at subtelomeric regions. In C. elegans, clk-2 mutants have elongated telomeres, and clk-2 overexpression can lead to telomere shortening. Tel2p has been reported to bind to telomeric DNA repeats in vitro. However, we find that a functional CLK-2::GFP fusion protein is cytoplasmic in worms. We discuss how the phenotype of clk-2 mutants could be the result of altered patterns of gene expression.

Amino Acid Sequence↗
PubMed2000

ROP-1, an RNA quality-control pathway component, affects Caenorhabditis elegans dauer formation.

Caenorhabditis elegans dauer formation is an alternative larval developmental pathway that the worm can take when environmental conditions become detrimental. Animals can survive several months in this stress-resistant stage and can resume normal development when growth conditions improve. Although the worms integrate a variety of sensory information to commit to dauer formation, it is currently unknown whether they also monitor internal cellular damage. The Ro ribonucleoprotein complex, which was initially described as a human autoantigen, is composed of one major 60-kDa protein, Ro60, that binds to one of four small RNA molecules, designated Y RNAs. Ro60 has been shown to bind mutant 5S rRNA molecules in Xenopus oocytes, suggesting a role for Ro60 in 5S rRNA biogenesis. Analysis of ribosomes from a C. elegans rop-1(-) strain, which is null for the expression of Ro60, demonstrated that they contain a high percentage of mutant 5S rRNA molecules, thereby strengthening the notion of a link between the rop-1 gene product and 5S rRNA quality control. The Ro particle was recently shown to be involved in the resistance of Deinococcus radiodurans to UV irradiation, suggesting a role for the Ro complex in stress resistance. We have studied the role of rop-1 in dauer formation. We present genetic and biochemical evidence that rop-1 interacts with dauer-formation genes and is involved in the regulation of the worms' entry into the dauer stage. Furthermore, we find that the rop-1 gene product undergoes a proteolytic processing step that is regulated by the dauer formation pathway via an aspartic proteinase. These results suggest that the Ro particle may function in an RNA quality-control checkpoint for dauer formation.

Animals↗
PubMed2000

clk-1, mitochondria, and physiological rates.

Mutations in the C. elegans maternal-effect gene clk-1 are highly pleiotropic, affecting the duration of diverse developmental and behavioral processes. They result in an average slowing of embryonic and post-embryonic development, adult rhythmic behaviors, reproduction, and aging.(1) CLK-1 is a highly conserved mitochondrial protein,(2,3) but even severe clk-1 mutations affect mitochondrial respiration only slightly.(3) Here, we review the evidence supporting the regulatory role of clk-1 in physiological timing. We also discuss possible models for the action of CLK-1, in particular, one proposing that CLK-1 is involved in the coordination of mitochondrial and nuclear function. BioEssays 22:48-56, 2000.

Amino Acid Sequence↗
PubMed1999

CLK-1 controls respiration, behavior and aging in the nematode Caenorhabditis elegans.

Mutations in the clk-1 gene of the nematode Caenorhabditis elegans result in an average slowing of a variety of developmental and physiological processes, including the cell cycle, embryogenesis, post-embryonic growth, rhythmic behaviors and aging. In yeast, a CLK-1 homologue is absolutely required for ubiquinone biosynthesis and thus respiration. Here we show that CLK-1 is fully active when fused to green fluorescent protein and is found in the mitochondria of all somatic cells. The activity of mutant mitochondria, however, is only very slightly impaired, as measured in vivo by a dye-uptake assay, and in vitro by the activity of succinate cytochrome c reductase. Overexpression of CLK-1 activity in wild-type worms can increase mitochondrial activity, accelerate behavioral rates during aging and shorten life span, indicating that clk-1 regulates and controls these processes. These observations also provide strong genetic evidence that mitochondria are causally involved in aging. Furthermore, the reduced respiration of the long-lived clk-1 mutants suggests that longevity is promoted by the age-dependent decrease in mitochondrial function that is observed in most species.

Aging↗
PubMed1999

Basolateral localization of the Caenorhabditis elegans epidermal growth factor receptor in epithelial cells by the PDZ protein LIN-10.

In Caenorhabditis elegans, the EGF receptor (encoded by let-23) is localized to the basolateral membrane domain of the epithelial vulval precursor cells, where it acts through a conserved Ras/MAP kinase signaling pathway to induce vulval differentiation. lin-10 acts in LET-23 receptor tyrosine kinase basolateral localization, because lin-10 mutations result in mislocalization of LET-23 to the apical membrane domain and cause a signaling defective (vulvaless) phenotype. We demonstrate that the previous molecular identification of lin-10 was incorrect, and we identify a new gene corresponding to the lin-10 genetic locus. lin-10 encodes a protein with regions of similarity to mammalian X11/mint proteins, containing a phosphotyrosine-binding and two PDZ domains. A nonsense lin-10 allele that truncates both PDZ domains only partially reduces lin-10 gene activity, suggesting that these protein interaction domains are not essential for LIN-10 function in vulval induction. Immunocytochemical experiments show that LIN-10 is expressed in vulval epithelial cells and in neurons. LIN-10 is present at low levels in the cytoplasm and at the plasma membrane and at high levels at or near the Golgi. LIN-10 may function in secretion of LET-23 to the basolateral membrane domain, or it may be involved in tethering LET-23 at the basolateral plasma membrane once it is secreted.

Amino Acid Sequence↗
PubMed1999

The levels of the RoRNP-associated Y RNA are dependent upon the presence of ROP-1, the Caenorhabditis elegans Ro60 protein.

The Ro ribonucleoproteins (RoRNP) consist of at least one major protein of 60 kD, Ro60, and one small associated RNA, designated Y RNA. Although RoRNP have been found in all vertebrate species examined so far, their function remains unknown. The Caenorhabditis elegans rop-1 gene previously has been identified as encoding a Ro60 homologue. We report here the phenotypic characterization of a C. elegans strain in which rop-1 has been disrupted. This is the first report regarding the inactivation of a major RoRNP constituent in any organism. The rop-1 mutant worms display no visible defects. However, at the molecular level, the disruption of rop-1 results in a dramatic decrease in the levels of the ROP-1-associated RNA (CeY RNA). Moreover, transgenic expression of wild-type rop-1 partially rescues the levels of CeY RNA. Considering that transgenes are poorly expressed in the germline, the fact that the rescue is only partial is most likely related to the high abundance of the CeY RNA in the adult germline and in embryos. The developmental expression pattern and localization of CeY RNA suggest a role for this molecule during embryogenesis. We conclude that, under laboratory culture conditions, ROP-1 does not play a crucial role in C. elegans.

Alleles↗
PubMed1999

Assessing the function of the Ro ribonucleoprotein complex using Caenorhabditis elegans as a biological tool.

The Ro ribonucleoprotein complex (Ro RNP) was initially described as an autoimmune target in human diseases such as systemic lupus erythematosus and Sjögren's syndrome. In Xenopus and human cells, its general structure is composed of one major protein of 60 kDa, Ro60, that binds to one of four small RNA molecules, designated Y RNAs. Although no function has been assigned to the Ro RNP, Ro60 has been shown to bind mutant 5S ribosomal RNA (rRNA) molecules in Xenopus oocytes, suggesting a role for Ro60 in 5S rRNA biogenesis. Ro60 has also been shown to participate in the regulation of the translational fate of the L4 ribosomal protein mRNA by interacting with the 5' untranslated region, again suggesting its possible implication in ribosome biogenesis. To identify the function of Ro RNP, we have taken a genetic approach in the nematode Caenorhabditis elegans. As such, we characterized the gene encoding the protein ROP-1, the homologue of the human Ro60 protein. Here, we review the phenotypic analysis of C. elegans rop-l(-) mutants and integrate these results into a model for the function of the Ro RNP particle.

Animals↗
PubMed1998

The genetics of caloric restriction in Caenorhabditis elegans.

Low caloric intake (caloric restriction) can lengthen the life span of a wide range of animals and possibly even of humans. To understand better how caloric restriction lengthens life span, we used genetic methods and criteria to investigate its mechanism of action in the nematode Caenorhabditis elegans. Mutations in many genes (eat genes) result in partial starvation of the worm by disrupting the function of the pharynx, the feeding organ. We found that most eat mutations significantly lengthen life span (by up to 50%). In C. elegans, mutations in a number of other genes that can extend life span have been found. Two genetically distinct mechanisms of life span extension are known: a mechanism involving genes that regulate dauer formation (age-1, daf-2, daf-16, and daf-28) and a mechanism involving genes that affect the rate of development and behavior (clk-1, clk-2, clk-3, and gro-1). We find that the long life of eat-2 mutants does not require the activity of DAF-16 and that eat-2; daf-2 double mutants live even longer than extremely long-lived daf-2 mutants. These findings demonstrate that food restriction lengthens life span by a mechanism distinct from that of dauer-formation mutants. In contrast, we find that food restriction does not further increase the life span of long-lived clk-1 mutants, suggesting that clk-1 and caloric restriction affect similar processes.

Aging↗
PubMed1998

Molecular genetics of life span in C. elegans: how much does it teach us?

Several loci have been identified in the nematode worm Caenorhabditis elegans that, when mutated, can increase life span. Three of these genes, age-1, daf-2 and clk-1, have now been cloned. Mutations in these three genes are highly pleiotropic and affect many aspects of worm development and behaviour, age-1 and daf-2 act in the same genetic pathway and have similar effects on the worm, age-1 encodes a homologue of the p110 subunit of phosphatidylinositol 3-kinase and daf-2 encodes an insulin receptor family member, clk-1 encodes a protein of unknown biochemical function similar to the yeast metabolic regulator Cat5p/Coq7p. The implications of these findings for our understanding of organismal ageing are discussed.

Amino Acid Sequence↗