Ruvkun, GB (2003) Harvey Lectures "The tiny RNA world."

Ruvkun, GB

[Harvey Lectures, 2003]

Horvitz HR (1988) Harvey Lect "Genetic control of Caenorhabditis elegans cell lineage."

Horvitz HR

[Harvey Lect, 1988]

Animal development involves a complex pattern of cell divisions ultimately leading to the generation of a highly diverse complement of cell types. What are the molecular mechanisms responsible for controlling patterns of cell division and for causing cells to become different from one another? To address the problem, my colleagues and I have been examining how genes control cell lineage and cell fate in the nematode Caenorhabditis elegans. The study of C. elegans was pioneered by Sydney Brenner (1973, 1974), who selected this organism because it is highly tractable genetically (e.g., see Herman, 1988)and because it is simple and essentially invariant in its cellular anatomy (the adult hermaphrodite is now known to contain a total of 959 somatic cell nuclei; Sulston and Horvitz, 1977; Kimble and Hirsh, 1979; Sulston et al., 1983). The invariance in C. elegans anatomy reflects a similar invariance in development. For example, the cell lineage of C. elegans is essentially the same in all individuals (Sulston and Horvitz, 1977; Kimble and Hirsh, 1979; Sulston et al., 1983). This cell lineage was determined by direct observation of living nematodes: individual nuclei were watched with the aid of Nomarski optics as they migrated,

Meyer BJ (1999) Harvey Lect "Sex and death of a worm: assessing and repressing X chromosomes."

Meyer BJ

[Harvey Lect, 1999]

A fundamental problem in biology is to determine how small quantitative differences in molecular signals are translated into alternative developmental fates and how developmental decisions are controlled by genetic regulatory hierarchies. A separate problem is to understand how gene expression is regulated at the level of whole chromosomes. Establishing the mechanistic link between higher-order chromatin structure and the regulation of gene expression is central to understanding this problem. These biological topics are addressed in this review through experiments that reveal the mechanisms by which the model organism Caenorhabditis elegans specifies its choice of sexual fate and regulates X chromosome-wide gene expression through the process of dosage compensation.

Harvey SC et al. (1997) International C. elegans Meeting "SEX DETERMINATION IN Strongyloides ratti."

Harvey SC, Viney ME

[International C. elegans Meeting, 1997]

Sex determination in S. ratti is not understood. S. ratti has two adult generations; one parasitic, parthenogenetic and female only and the other free-living, dioecious and sexual. The progeny of the parasitic females have the potential to develop into both males and females, whereas the progeny of the free-living sexual generation all develop into females. Cyological studies by other workers suggested that sex determination was an XX/XO system, however other cytological studies of S. ratti have [previously] produced wrong conclusions. Further, the XX/X0 system does not explain the production of only female progeny by the free-living adult generation. To investiagte these problems, we have isolated putative sex-linked DNA fragments. This has been done by screening anonymous genomic DNA fragments against dot blots of free-living male and female DNA. Semi-quantitative PCR was then used to further confirm this quantitative difference between free-living males and females and then to screen other life-cycle stages.

Harvey, A. et al. (2019) International Worm Meeting "Using fast-acting temperature-sensitive alleles to identify genes required for meiosis I anaphase."

Harvey, A., Bowerman, B.

[International Worm Meeting, 2019]

During oogenesis, the maternal genome is replicated, recombined, and then partitioned via two rounds of meiotic division to produce haploid gametes. Each meiotic division depends on a highly dynamic microtubule-based spindle structure to organize and, during anaphase, segregate chromosomes; errors in this process can result in defects such as aneuploidy, infertility, and miscarriages. However, the genetic requirements for chromosome segregation during meiosis I anaphase are poorly understood, due in part to gene requirements earlier during spindle assembly which makes assessment of their later roles during anaphase difficult. Furthermore, because from beginning to end anaphase lasts only about 5 minutes, methods such as RNAi or degron-mediated gene knockdowns lack the temporal resolution needed to inactivate genes specifically during anaphase. To circumvent these limitations, we are using temperature-sensitive (TS) mutants to directly assess gene requirements during meiosis I anaphase, upshifting fast-acting TS alleles to a restrictive temperature with a CherryTemp live imaging stage that allows precise temperature control of mounted animals. Our initial experiments have focused on the XMAP215 homolog ZYG-9. From these experiments we have found that upshift of zyg-9(or623ts)at metaphase causes reversion of the bipolar spindle back into a multipolar-like state, suggesting a requirement for ZYG-9 in maintaining pole stability. Eventually, these spindles recover, reestablishing bipolarity as meiosis progresses and ultimately segregating two chromosome masses at the end of meiosis I. These preliminary results suggest that ZYG-9 is required post-metaphase for spindle pole stability. Our ongoing investigation aims to further define ZYG-9 requirements before and beyond metaphase at high temporal resolution. We also are assessing requirements for the kinesin-12 family member protein KLP-18, with preliminary data indicating that KLP-18 is required for chromosome segregation during anaphase. Our goal is to identify genes that control spindle dynamics during anaphase to ensure faithful chromosome segregation in developing oocytes.

Harvey SC et al. (2000) Proc Biol Sci "The control of morph development in the parasitic nematode Strongyloides ratti."

Viney ME, Gemmill AW, Harvey SC, Read AF

[Proc Biol Sci, 2000]

The parasitic nematode Strongyloides ratti has a complex life cycle. The progeny of the parasitic females can develop into three distinct morphs, namely directly developing infective third-stage larvae (iL3s), free-living adult males and free-living adult females. We have analysed of the effect of host immune status (an intra-host factor), environmental temperature (an extra-host factor) and their interaction on the proportion of larvae that develop into these three morphs. The results are consistent with the developmental decision of larvae being controlled by at least two discrete developmental switches. One is a sex-determination event that is affected by host immune status and the other is a switch between alternative female morphs that is affected by both host immune status and environmental temperature. These findings clarify the basis of the life cycle of S. ratti and demonstrate how such complex life cycles can result from a combination of simple developmental switches.

Harvey, L-M. et al. (2019) International Worm Meeting "Identification and characterization of new actors of the RNAi pathway."

Harvey, L-M., Simard, M.J., Kakumani, P.K.

[International Worm Meeting, 2019]

The RNA interference pathway (RNAi) uses either ectopic (exo-RNAi) or endogenous (endo-RNAi) short non-coding RNA molecules known as short interfering RNA (siRNA) to regulate gene expression. To this day, knowledge on this regulation pathway was mainly obtained with studies made in plants and nematodes where it has been described as important for the genome protection and stability against pathogens. In contrast, our understanding of this regulatory pathway in higher eukaryotes is underwhelming. In mammals, a strong presence of these endogenous siRNA (endo-siRNA) has been detected in embryonic stem cells (ESCs), but in opposition to plants and nematodes, their function is yet to be fully understood. In an effort to address this question, we recently identified specific proteins associated to the short interfering RNA induced silencing complex (siRISC) in mouse ESCs using a 2'-O-methyl pulldown of an endo-siRNA followed by mass spectrometry. Beside Argonaute proteins, we identified 10 proteins associated with the mouse siRNA that are also encoded in the C. elegans genome, suggesting that they may play a conserved role in the endo-siRNA pathway. Therefore, we initiated the characterization of these new siRNA interactors in C. elegans, a biological system in which the tools and reagents needed to study components of the endo-siRNA pathway are well established in contrast to mammalian cells. Several molecular and genetic approaches are currently used to quickly understand the implication of these new factors in the RNAi pathway. During this meeting, we will present our recent progress on the characterization of some of the identified proteins. At term, this characterization will contribute to define the role of new conserved actors in the endo-siRNA pathway and lead to a better understanding of the pathway in mammals.

Harvey SC et al. (2000) European Worm Meeting "The control of morph development in the parasitic nematode Strongyloides ratti"

Harvey SC, Viney ME

[European Worm Meeting, 2000]

The life-cycle of the parasitic nematode Strongyloides ratti is complex. The progeny of the parasitic females can develop into three distinct morphs, namely directly developing infective third stage larvae (iL3s), free-living adult males and free-living adult females. We have analysed of the effect of host immune status (an intra-host factor) and environmental temperature (an extra-host factor) and their interaction on the proportion of larvae that develop into these three morphs. The results are consistent with the developmental decision of larvae being controlled by at least two discrete developmental switches. One developmental decision is a sex determination event and other is a switch between alternative female morphs. These findings clarify the basis of the life-cycle of S. ratti and demonstrate how such complex life-cycles can result from a combination of simple developmental switches.

S.C. Harvey et al. (2006) European Worm Meeting "The Genetics of Natural Variation in the Phenotypic Plasticity of Dauer Larvae Development"

A. Shorto, S.C. Harvey, M.E. Viney

[European Worm Meeting, 2006]

S.C. Harvey, A. Shorto and M.E. Viney . Natural isolates (including N2 and DR1350) of the free-living nematode Caenorhabditis elegans vary in their phenotypic plasticity of dauer larvae development. For example, some lines appear to be highly sensitive to dauer inducing conditions while others are less so. We have sought to investigate the causes and consequences of this plasticity.. To determine the genetic basis of this plasticity of dauer development we have undertaken a quantitative trait loci (QTL) mapping based analysis of recombinant inbred lines (RILs) produced from crosses between N2 x DR1350. This has identified several regions containing candidate QTLs that affect the plasticity of dauer development. Nearly isogenic lines (NILs) have been constructed for a candidate QTL on chromosome II, with analysis of these NILs confirming that the region contains genes that affect the plasticity of dauer larvae development. This nearly isogenic region contains a maximum of 441 genes, none of which have previously been identified as being part of the genetic pathway regulating dauer development. Overall, these data have developed a clearer picture of the genetics underlying natural variation in a complex trait and demonstrate that the analysis of natural variation can reveal genes not identified by other genetic approaches.

Harvey SC et al. (2009) BMC Genomics "Natural variation in gene expression in the early development of dauer larvae of Caenorhabditis elegans."

Shorto A, Barker GL, Viney ME, Harvey SC

[BMC Genomics, 2009]

BACKGROUND: The free-living nematode Caenorhabditis elegans makes a developmental decision based on environmental conditions: larvae either arrest as dauer larva, or continue development into reproductive adults. There is natural variation among C. elegans lines in the sensitivity of this decision to environmental conditions; that is, there is variation in the phenotypic plasticity of dauer larva development. We hypothesised that these differences may be transcriptionally controlled in early stage larvae. We investigated this by microarray analysis of different C. elegans lines under different environmental conditions, specifically the presence and absence of dauer larva-inducing pheromone. RESULTS: There were substantial transcriptional differences between four C. elegans lines under the same environmental conditions. The expression of approximately 2,000 genes differed between genetically different lines, with each line showing a largely line-specific transcriptional profile. The expression of genes that are markers of larval moulting suggested that the lines may be developing at different rates. The expression of a total of 89 genes was putatively affected by dauer larva or non-dauer larva-inducing conditions. Among the upstream regions of these genes there was an over-representation of DAF-16-binding motifs. CONCLUSION: Under the same environmental conditions genetically different lines of C. elegans had substantial transcriptional differences. This variation may be due to differences in the developmental rates of the lines. Different environmental conditions had a rather smaller effect on transcription. The preponderance of DAF-16-binding motifs upstream of these genes was consistent with these genes playing a key role in the decision between development into dauer or into non-dauer larvae. There was little overlap between the genes whose expression was affected by environmental conditions and previously identified loci involved in the plasticity of dauer larva development.