- Email: [email protected]
Developmental Genetics of Caenorhabditis elegans
Developmental Genetics of Caenorhabditis elegans E Kage-Nakadai and S Mitani, Tokyo Women’s Medical University School of Medicine, Tokyo, Japan
© 2013 Elsevier Inc. All rights reserved.
This article is a revision of the previous edition article by J Hodgkin, volume 1, pp 531–532, © 2001, Elsevier Inc.
Glossary Dauer An alternative developmental state in which the larva arrests development, allowing it to survive harsh conditions. Dicer A ribonuclease involved in the production of microRNAs and small interfering RNAs (siRNAs). Differential interference contrast microscopy Microscopy (also known as Nomarski microscopy) by which the height difference of a specimen that is invisible with bright field becomes a relief-like or three-dimensional image with improved contrast.
Introduction In 1965, Brenner established the nematode Caenorhabditis elegans as a model organism, and it is now one of the simplest and most powerful tools available for genetic dissection in developmental biology. C. elegans is a nematode of about 1 mm in length, with two sexes, hermaphrodite and male. Hermaphrodites can reproduce by both self-fertilization and mating with males. C. elegans develops through four larval stages and matures within approximately 3–4 days. C. elegans has a simple body plan with an outer tube consisting of a cuticle, hypodermis (epidermis), neurons, and muscles sur rounding a pseudocoelomic space that contains the intestine and the gonad. The adult hermaphrodite has only 959 somatic cells and the adult male has only 1031, all lineages of which have been described by Sulston and Horvitz. C. elegans is transparent, which allowed Chalfie to apply the green fluores cent protein that is now widely used in many species throughout biology. These favorable features offer great poten tial for developmental genetics.
Distal tip cell (DTC) A somatic cell located at the distal tip of each gonad arm. Hermaphrodite An animal possessing both male and female reproductive organs. miRNA Noncoding small RNA. Spermatheca An enlarged portion of the hermaphrodite gonad between the oviduct and the uterus that stores sperm. Vulva The external female (hermaphrodite in Caenorhabditis elegans) organ through which the sperm enter and fertilized eggs leave the gonad.
cells, encodes a Notch family receptor. The lag-2 gene, expressed in the DTC, encodes a protein homologous to Drosophila Delta, a Notch ligand.
Cell Division, Polarity, and Cell Fate Specification in the Early Embryo In C. elegans, embryogenesis occurs within a solute-impermeable eggshell that is transparent, enabling observations of both cell cleavage and cell differentiation in the living embryo with both light and fluorescence microscopy. A newly fertilized egg under goes the first few divisions within a couple of hours, during which the polarity of the embryo is established. Several maternal-effect lethal mutations have led to the identification of genes involved in establishing early embryonic polarity, for example, mutations in the par (partitioning-defective) genes affect the first division, leading to the disruption of anterior– posterior polarity.
Programmed Cell Death
Genetic Dissection of C. elegans Development Germline Development The C. elegans hermaphrodite gonad comprises two U-shaped tubular arms, each terminating proximally at a spermatheca, which is discussed elsewhere in this encyclopedia. The germ cells undergo a sequential process of proliferation, meiotic pro phase progression, and gametogenesis along the distal to proximal direction in the gonadal tube. The distal end of the gonad arms is composed of the somatic distal tip cells (DTCs). Cell ablation analyses revealed that the DTCs promote germ cell proliferation and inhibit entry into meiosis. The screening of mutations with sterile (Ste) phenotypes has identified many genes with roles in germline development. These genes are involved in germline specification and survival, sex determina tion, oogenesis, and spermatogenesis. In addition, some of the genes (e.g., glp-1 and lag-2) act as signaling molecules from the DTC to the germ cells. The glp-1 gene, which is required in germ
308
In C. elegans, programmed cell death is highly reproducible, but not essential for viability. Cells dying due to programmed cell death undergo morphologic changes that can be observed in living animals using differential interference contrast micro scopy. Of the 1090 cells generated during hermaphrodite somatic development, 131 undergo programmed cell death. Most somatic cell death occurs during embryogenesis. In males, a few cells die in the fourth larval stage, facilitating the formation of the male-specific organs. Genetic screens have identified over 100 mutations that affect programmed cell death in C. elegans. Cloning and characterization of these genes revealed a programmed cell death or apoptotic pathway that is conserved from C. elegans to human. The egl-1 gene encodes a protein with a Bcl-2 homology domain-3 that is commonly found in proapoptotic members of the Bcl-2 family. The ced-3 gene encodes a protein which is member of cysteine proteases (caspases) family. The ced-4 gene encodes an ortholog of human Apaf-1. The ced-9 gene encodes a protein homologous to the human anti-apoptotic Bcl-2 family.
Brenner’s Encyclopedia of Genetics, 2nd Edition, Volume 2
doi:10.1016/B978-0-12-374984-0.00396-X
Developmental Genetics of Caenorhabditis elegans
Sex Determination C. elegans hermaphrodites have two X chromosomes (XX) and males have one (XO). The small twofold difference in the X chromosome number specifies not only the sexual characteris tics of the adult but also all somatic cells throughout development, starting at the 24- to 28-cell stage. Many of the genes that determine sexual fate have been identified by genetic screens of mutations that cause phenotypes such as transformer of sex (Tra), hermaphroditization (Her), feminization (Fem), sex and dosage compensation defects (Sdc), and XO-lethal (Xol). Comparison between C. elegans and Drosophila or mam mals, however, reveals that there are only a few molecular overlaps in sex determination, although there are some simila rities in the sex determination strategies.
Vulval Development The hermaphrodite vulva contains only 22 cells. Vulval deve lopment processes are divided into essentially four stages: (1) the vulval precursor cells are born; (2) cell–cell interactions specify the fate of the vulval precursor cells; (3) the vulval precursor cells execute their predetermined fates, producing the appropriate number and types of vulval cells; and (4) the vulval cells undergo cell movement, fusion, and evasion to form the mature vulva. The simplicity and invariance of the vulval formation pattern, and the fact that vulva formation is not essential for viability, led to the development of a satura tion screen for mutants with abnormal vulva formation, such as ‘vulvaless’ mutants or ‘multivulva’ mutants. These types of genetic screens for mutations affecting vulval development and suppressors of extant vulval mutants led to the identification of central players in signal transduction, which are highly con served across animal phylogeny. Genetic screens and epistasis analyses led to the discovery of the signaling pathways of LET 23/EGFR-LET-60/Ras-MAP kinase, LIN-12/Notch, and WNT.
Muscle Development C. elegans contains several groups of muscles with diverse func tion such as locomotion, pharyngeal pumping, intestinal contraction, and egg laying. The body wall muscles are used for locomotion. Many mutants in which locomotion is affected have been isolated, leading to the identification of genes encoding major components of the muscle structure. Brenner’s screen for uncoordinated (Unc) mutants identified the unc-54 gene, which encodes one of the myosin heavy-chain isoforms. Actin mutants were first identified as semidominant, paralyzed animals desig nated unc-92. The results of genetic studies of the C. elegans MyoD homolog, encoded by the single gene hlh-1, are paradoxical. Mutational analysis indicates that, unlike vertebrate MyoD, HLH-1 is not the only factor that executes myogenesis. HLH-1 can function in vertebrate tissue culture to promote muscle dif ferentiation, however, and at least one of the vertebrate MyoD family members can substitute for C. elegans hlh-1.
Nervous System Development The C. elegans nervous system contains 302 neurons. Neurons are generated by invariant patterns of cell division and migra tion and are interconnected to form neural circuits by regulated
309
axonal growth cone migration and synaptic connections. Genes that define cell lineage and neural identity were detected in genetic screens for mutants with loss of cell lineages (Lin) or with defects of neural function such as locomotion (Unc) and mechanosensation (Mec). Many of these genes encode tran scription factors. lin-32, encoding a helix–loop–helix transcription factor, and unc-86 and mec-3, encoding homeo domain transcription factors, commit cell lineages of sensory neurons. unc-4, which encodes a Pax-related transcription fac tor, is necessary for the specification of a class of motor neurons. Loss-of-function mutations in unc-4 lead to backward movement defects, but have little effect on forward movement. UNC-4 defines synaptic specificity indirectly by regulating the expression of downstream genes. Many of the genes that regulate axon guidance were originally identified among a large collection of unc mutations. UNC-6/Netrin guidance cue and UNC-40/DCC and UNC-5 Netrin receptors mediate cell and growth cone migrations along the dorsal–ventral axis in C. elegans. On the other hand, reverse genetic approaches revealed that SLT-1/Slit acts in midline, dorsal–ventral, and anterior–posterior guidance via the SAX-3/Robo receptor.
Heterochronic Genes A change in the relative timing of developmental events is called ‘heterochrony’ (khronos in Greek, meaning ‘time’). Mutations that affect the timing of stage-specific events have been studied intensively. A nuclear protein encoded by lin-14 was identified as a heterochronic gene required for the regulation of the larval transition in the timing of the decrease in the LIN-14 protein level. Genetic and molecular experiments revealed that lin-4 negatively regulates lin-14. Cloning of lin-4 revealed that it does not encode a protein, but rather it encodes a noncoding small RNA, called microRNA (miRNA), that contains sequences com plementary to an element in the 3′-untranslated region of lin-14 mRNA. lin-4 miRNA downregulates the expression of lin-14 and other target genes that regulate developmental timing through interacting with partially complementary sequences in the 3′ -untranslated regions of the target genes. The identification of lin-4 and lin-14 set a precedent for the study of the second miRNA gene, let-7, and its target mRNA, lin-41. let-7 is tran scribed into a precursor RNA of approximately 70 nucleotides, which is processed into a mature miRNA of 22 nucleotides by the RNase III, Dicer (DCR-1 in C. elegans). Soon after the dis covery of let-7, let-7 orthologs were discovered in fly and human. The let-7 family and other miRNAs are currently thought to be involved in stem cell differentiation and cancer.
Dauer Development Under conditions of increased population density and limited food, C. elegans arrests development at the second molt and undergoes morphologically and behaviorally specialized alterations to enhance dispersal and long-term survival (dauern in German, meaning ‘to last’). Dauer larvae survive 4–8 times longer than the normal 2- to 3-week life span of C. elegans. The dauer state itself is thought to be one of nonaging because the duration of the dauer stage does not affect the post-dauer life span. Genetic analysis of dauer-constitutive mutants identified daf-23 (now referred to as age-1) and daf-2, and the analysis of dauer-defective mutants identified daf-16. Reduced daf-2 and
310
Developmental Genetics of Caenorhabditis elegans
age-1 gene activities dramatically extend the life span. Moreover, daf-16 is required for the extended life span of daf-2 and age-1 mutants. Several years after these discoveries, researchers demonstrated that daf-2 encodes an insulin receptor homolog, age-1 encodes phosphoinositide 3-kinase, and daf-16 encodes the forkhead box class O transcriptional factor. Insulin signaling pathways and forkhead box class O transcription factors are also involved in the life span of flies and mice.
Conclusions and Future Prospects Genetic screens for mutations that affect the developmental pro cesses of C. elegans are a powerful approach to exploit the genes or pathways underlying the conserved animal developmental sys tems. In some cases, the impact of the discoveries has not been limited to developmental biology and has greatly expanded to new research fields and paradigms. Over recent years, deep sequencing technology, which was first applied to the C. elegans genome, has changed the field of genetics. Whole-genome sequences of many organisms have now been disclosed, and forward and reverse genetics are indispensable to biologic studies. Currently, the National Bioresource Project in Japan and the C. elegans Gene Knockout Consortium have isolated over 6000 deletion mutants. The collection continues to grow, offering more opportunities to validate and deepen the understanding of genetic screens. Developmental genetic studies of C. elegans will contribute to new and exciting discoveries in the coming decade.
See also: Caenorhabditis elegans (Nematode); Cell Division in Caenorhabditis elegans; Embryonic Development of the Nematode Caenorhabditis elegans; Neurogenetics of neuro transmitter Release in Caenorhabditis elegans; Oogenesis, in Caenorhabditis elegans; Spermatogenesis in C. elegans.
Further Reading Conradt B and Xue D (2005) Programmed cell death. WormBook 1–13.
Hu PJ (2007) Dauer. WormBook 1–19.
Kenyon C (2011) The first long-lived mutants: Discovery of the insulin/IGF-1 pathway for
ageing. Philosophical Transactions of the Royal Society London: Series B, Biological Sciences 366: 9–16. Riddle D, Blumenthal T, Meyer B, and Priess J (eds.) (1997) C. elegans II. New York: Cold Spring Harbor Laboratory Press. Ruvkun G (2008) The perfect storm of tiny RNAs. Nature Medicine 14: 1041–1045. Sternberg PW (2005) Vulval development. WormBook 1–28. Vella MC and Slack FJ (2005) C. elegans microRNAs. WormBook 1–9. Wood WB (ed.) (1998) The Nematode Caenorhabditis elegans. New York: Cold Spring Harbor Laboratory Press.
Relevant Websites http://www.cbs.umn.edu – Caenorhabditis Genetics Center (CGC).
http://www.shigen.nig.ac.jp – National Bioresource Project (NBRP).
http://www.wormatlas.org – WormAtlas.
http://www.wormbase.org – WormBase.
© 2013 Elsevier Inc. All rights reserved.
This article is a revision of the previous edition article by J Hodgkin, volume 1, pp 531–532, © 2001, Elsevier Inc.
Glossary Dauer An alternative developmental state in which the larva arrests development, allowing it to survive harsh conditions. Dicer A ribonuclease involved in the production of microRNAs and small interfering RNAs (siRNAs). Differential interference contrast microscopy Microscopy (also known as Nomarski microscopy) by which the height difference of a specimen that is invisible with bright field becomes a relief-like or three-dimensional image with improved contrast.
Introduction In 1965, Brenner established the nematode Caenorhabditis elegans as a model organism, and it is now one of the simplest and most powerful tools available for genetic dissection in developmental biology. C. elegans is a nematode of about 1 mm in length, with two sexes, hermaphrodite and male. Hermaphrodites can reproduce by both self-fertilization and mating with males. C. elegans develops through four larval stages and matures within approximately 3–4 days. C. elegans has a simple body plan with an outer tube consisting of a cuticle, hypodermis (epidermis), neurons, and muscles sur rounding a pseudocoelomic space that contains the intestine and the gonad. The adult hermaphrodite has only 959 somatic cells and the adult male has only 1031, all lineages of which have been described by Sulston and Horvitz. C. elegans is transparent, which allowed Chalfie to apply the green fluores cent protein that is now widely used in many species throughout biology. These favorable features offer great poten tial for developmental genetics.
Distal tip cell (DTC) A somatic cell located at the distal tip of each gonad arm. Hermaphrodite An animal possessing both male and female reproductive organs. miRNA Noncoding small RNA. Spermatheca An enlarged portion of the hermaphrodite gonad between the oviduct and the uterus that stores sperm. Vulva The external female (hermaphrodite in Caenorhabditis elegans) organ through which the sperm enter and fertilized eggs leave the gonad.
cells, encodes a Notch family receptor. The lag-2 gene, expressed in the DTC, encodes a protein homologous to Drosophila Delta, a Notch ligand.
Cell Division, Polarity, and Cell Fate Specification in the Early Embryo In C. elegans, embryogenesis occurs within a solute-impermeable eggshell that is transparent, enabling observations of both cell cleavage and cell differentiation in the living embryo with both light and fluorescence microscopy. A newly fertilized egg under goes the first few divisions within a couple of hours, during which the polarity of the embryo is established. Several maternal-effect lethal mutations have led to the identification of genes involved in establishing early embryonic polarity, for example, mutations in the par (partitioning-defective) genes affect the first division, leading to the disruption of anterior– posterior polarity.
Programmed Cell Death
Genetic Dissection of C. elegans Development Germline Development The C. elegans hermaphrodite gonad comprises two U-shaped tubular arms, each terminating proximally at a spermatheca, which is discussed elsewhere in this encyclopedia. The germ cells undergo a sequential process of proliferation, meiotic pro phase progression, and gametogenesis along the distal to proximal direction in the gonadal tube. The distal end of the gonad arms is composed of the somatic distal tip cells (DTCs). Cell ablation analyses revealed that the DTCs promote germ cell proliferation and inhibit entry into meiosis. The screening of mutations with sterile (Ste) phenotypes has identified many genes with roles in germline development. These genes are involved in germline specification and survival, sex determina tion, oogenesis, and spermatogenesis. In addition, some of the genes (e.g., glp-1 and lag-2) act as signaling molecules from the DTC to the germ cells. The glp-1 gene, which is required in germ
308
In C. elegans, programmed cell death is highly reproducible, but not essential for viability. Cells dying due to programmed cell death undergo morphologic changes that can be observed in living animals using differential interference contrast micro scopy. Of the 1090 cells generated during hermaphrodite somatic development, 131 undergo programmed cell death. Most somatic cell death occurs during embryogenesis. In males, a few cells die in the fourth larval stage, facilitating the formation of the male-specific organs. Genetic screens have identified over 100 mutations that affect programmed cell death in C. elegans. Cloning and characterization of these genes revealed a programmed cell death or apoptotic pathway that is conserved from C. elegans to human. The egl-1 gene encodes a protein with a Bcl-2 homology domain-3 that is commonly found in proapoptotic members of the Bcl-2 family. The ced-3 gene encodes a protein which is member of cysteine proteases (caspases) family. The ced-4 gene encodes an ortholog of human Apaf-1. The ced-9 gene encodes a protein homologous to the human anti-apoptotic Bcl-2 family.
Brenner’s Encyclopedia of Genetics, 2nd Edition, Volume 2
doi:10.1016/B978-0-12-374984-0.00396-X
Developmental Genetics of Caenorhabditis elegans
Sex Determination C. elegans hermaphrodites have two X chromosomes (XX) and males have one (XO). The small twofold difference in the X chromosome number specifies not only the sexual characteris tics of the adult but also all somatic cells throughout development, starting at the 24- to 28-cell stage. Many of the genes that determine sexual fate have been identified by genetic screens of mutations that cause phenotypes such as transformer of sex (Tra), hermaphroditization (Her), feminization (Fem), sex and dosage compensation defects (Sdc), and XO-lethal (Xol). Comparison between C. elegans and Drosophila or mam mals, however, reveals that there are only a few molecular overlaps in sex determination, although there are some simila rities in the sex determination strategies.
Vulval Development The hermaphrodite vulva contains only 22 cells. Vulval deve lopment processes are divided into essentially four stages: (1) the vulval precursor cells are born; (2) cell–cell interactions specify the fate of the vulval precursor cells; (3) the vulval precursor cells execute their predetermined fates, producing the appropriate number and types of vulval cells; and (4) the vulval cells undergo cell movement, fusion, and evasion to form the mature vulva. The simplicity and invariance of the vulval formation pattern, and the fact that vulva formation is not essential for viability, led to the development of a satura tion screen for mutants with abnormal vulva formation, such as ‘vulvaless’ mutants or ‘multivulva’ mutants. These types of genetic screens for mutations affecting vulval development and suppressors of extant vulval mutants led to the identification of central players in signal transduction, which are highly con served across animal phylogeny. Genetic screens and epistasis analyses led to the discovery of the signaling pathways of LET 23/EGFR-LET-60/Ras-MAP kinase, LIN-12/Notch, and WNT.
Muscle Development C. elegans contains several groups of muscles with diverse func tion such as locomotion, pharyngeal pumping, intestinal contraction, and egg laying. The body wall muscles are used for locomotion. Many mutants in which locomotion is affected have been isolated, leading to the identification of genes encoding major components of the muscle structure. Brenner’s screen for uncoordinated (Unc) mutants identified the unc-54 gene, which encodes one of the myosin heavy-chain isoforms. Actin mutants were first identified as semidominant, paralyzed animals desig nated unc-92. The results of genetic studies of the C. elegans MyoD homolog, encoded by the single gene hlh-1, are paradoxical. Mutational analysis indicates that, unlike vertebrate MyoD, HLH-1 is not the only factor that executes myogenesis. HLH-1 can function in vertebrate tissue culture to promote muscle dif ferentiation, however, and at least one of the vertebrate MyoD family members can substitute for C. elegans hlh-1.
Nervous System Development The C. elegans nervous system contains 302 neurons. Neurons are generated by invariant patterns of cell division and migra tion and are interconnected to form neural circuits by regulated
309
axonal growth cone migration and synaptic connections. Genes that define cell lineage and neural identity were detected in genetic screens for mutants with loss of cell lineages (Lin) or with defects of neural function such as locomotion (Unc) and mechanosensation (Mec). Many of these genes encode tran scription factors. lin-32, encoding a helix–loop–helix transcription factor, and unc-86 and mec-3, encoding homeo domain transcription factors, commit cell lineages of sensory neurons. unc-4, which encodes a Pax-related transcription fac tor, is necessary for the specification of a class of motor neurons. Loss-of-function mutations in unc-4 lead to backward movement defects, but have little effect on forward movement. UNC-4 defines synaptic specificity indirectly by regulating the expression of downstream genes. Many of the genes that regulate axon guidance were originally identified among a large collection of unc mutations. UNC-6/Netrin guidance cue and UNC-40/DCC and UNC-5 Netrin receptors mediate cell and growth cone migrations along the dorsal–ventral axis in C. elegans. On the other hand, reverse genetic approaches revealed that SLT-1/Slit acts in midline, dorsal–ventral, and anterior–posterior guidance via the SAX-3/Robo receptor.
Heterochronic Genes A change in the relative timing of developmental events is called ‘heterochrony’ (khronos in Greek, meaning ‘time’). Mutations that affect the timing of stage-specific events have been studied intensively. A nuclear protein encoded by lin-14 was identified as a heterochronic gene required for the regulation of the larval transition in the timing of the decrease in the LIN-14 protein level. Genetic and molecular experiments revealed that lin-4 negatively regulates lin-14. Cloning of lin-4 revealed that it does not encode a protein, but rather it encodes a noncoding small RNA, called microRNA (miRNA), that contains sequences com plementary to an element in the 3′-untranslated region of lin-14 mRNA. lin-4 miRNA downregulates the expression of lin-14 and other target genes that regulate developmental timing through interacting with partially complementary sequences in the 3′ -untranslated regions of the target genes. The identification of lin-4 and lin-14 set a precedent for the study of the second miRNA gene, let-7, and its target mRNA, lin-41. let-7 is tran scribed into a precursor RNA of approximately 70 nucleotides, which is processed into a mature miRNA of 22 nucleotides by the RNase III, Dicer (DCR-1 in C. elegans). Soon after the dis covery of let-7, let-7 orthologs were discovered in fly and human. The let-7 family and other miRNAs are currently thought to be involved in stem cell differentiation and cancer.
Dauer Development Under conditions of increased population density and limited food, C. elegans arrests development at the second molt and undergoes morphologically and behaviorally specialized alterations to enhance dispersal and long-term survival (dauern in German, meaning ‘to last’). Dauer larvae survive 4–8 times longer than the normal 2- to 3-week life span of C. elegans. The dauer state itself is thought to be one of nonaging because the duration of the dauer stage does not affect the post-dauer life span. Genetic analysis of dauer-constitutive mutants identified daf-23 (now referred to as age-1) and daf-2, and the analysis of dauer-defective mutants identified daf-16. Reduced daf-2 and
310
Developmental Genetics of Caenorhabditis elegans
age-1 gene activities dramatically extend the life span. Moreover, daf-16 is required for the extended life span of daf-2 and age-1 mutants. Several years after these discoveries, researchers demonstrated that daf-2 encodes an insulin receptor homolog, age-1 encodes phosphoinositide 3-kinase, and daf-16 encodes the forkhead box class O transcriptional factor. Insulin signaling pathways and forkhead box class O transcription factors are also involved in the life span of flies and mice.
Conclusions and Future Prospects Genetic screens for mutations that affect the developmental pro cesses of C. elegans are a powerful approach to exploit the genes or pathways underlying the conserved animal developmental sys tems. In some cases, the impact of the discoveries has not been limited to developmental biology and has greatly expanded to new research fields and paradigms. Over recent years, deep sequencing technology, which was first applied to the C. elegans genome, has changed the field of genetics. Whole-genome sequences of many organisms have now been disclosed, and forward and reverse genetics are indispensable to biologic studies. Currently, the National Bioresource Project in Japan and the C. elegans Gene Knockout Consortium have isolated over 6000 deletion mutants. The collection continues to grow, offering more opportunities to validate and deepen the understanding of genetic screens. Developmental genetic studies of C. elegans will contribute to new and exciting discoveries in the coming decade.
See also: Caenorhabditis elegans (Nematode); Cell Division in Caenorhabditis elegans; Embryonic Development of the Nematode Caenorhabditis elegans; Neurogenetics of neuro transmitter Release in Caenorhabditis elegans; Oogenesis, in Caenorhabditis elegans; Spermatogenesis in C. elegans.
Further Reading Conradt B and Xue D (2005) Programmed cell death. WormBook 1–13.
Hu PJ (2007) Dauer. WormBook 1–19.
Kenyon C (2011) The first long-lived mutants: Discovery of the insulin/IGF-1 pathway for
ageing. Philosophical Transactions of the Royal Society London: Series B, Biological Sciences 366: 9–16. Riddle D, Blumenthal T, Meyer B, and Priess J (eds.) (1997) C. elegans II. New York: Cold Spring Harbor Laboratory Press. Ruvkun G (2008) The perfect storm of tiny RNAs. Nature Medicine 14: 1041–1045. Sternberg PW (2005) Vulval development. WormBook 1–28. Vella MC and Slack FJ (2005) C. elegans microRNAs. WormBook 1–9. Wood WB (ed.) (1998) The Nematode Caenorhabditis elegans. New York: Cold Spring Harbor Laboratory Press.
Relevant Websites http://www.cbs.umn.edu – Caenorhabditis Genetics Center (CGC).
http://www.shigen.nig.ac.jp – National Bioresource Project (NBRP).
http://www.wormatlas.org – WormAtlas.
http://www.wormbase.org – WormBase.