Phillips PC (1995) International C. elegans Meeting "DISTINGUISHING CHAOS FROM NOISE IN NEMATODE POPULATION DYNAMICS"

Phillips PC

[International C. elegans Meeting, 1995]

C. elegans populations raised via serial transfer with discrete generations display fluctuations in population size that can span several orders of magnitude. This is somewhat unusual since the worms are maintained under controlled conditions on a fixed amount of resource (OP50). Large fluctuations in population size in an apparently constant environment could be the signature of deterministic chaos induced by density dependent population size regulation and the very large reproductive capacity of C. elegans. To test this hypothesis, I raised four replicate lines of worms for 100 generations at 20 C, transferring the populations every four days using the hypochlorite cleaning procedure (i.e., only eggs were transferred). Analysis of the resulting time series shows that there is strong density dependent regulation in these populations, but that the size fluctuations are driven mostly by environmental variation rather than deterministic chaos. The environmental influences are probably caused by variation in the age and quality of E. coli cultures and plates, as well as small differences in the timing of transfers. Large scale sensitivity resulting from small amounts of input variation probably arises from "local chaos" during different parts of the population regulation cycle. This study demonstrates that C. elegans can be developed into a useful model system for ecological and evolutionary studies.

PC Phillips et al. (1999) International C. elegans Meeting "Influence of temperature on reproductive rate in C. elegans: r versus Ro"

BL Armstrong, RB Huey, CP Coucke, PC Phillips

[International C. elegans Meeting, 1999]

Temperature is one of the primary determinants of ectotherm performance and fitness. Because of its quick generation time, easy maintenance, and powerful genetics, C. elegans provides an ideal model for testing many of the central theories of thermal ecology. As a precursor to a general set of experiments in this area, we have been characterizing thermal performance curves for C. elegans (N2) over a wide range of temperatures. There are potentially many ecologically relevant measures of performance. Two standard measures of fitness, the net reproductive rate ( R o ) and the intrinsic rate of increase ( r ), differ in their sensitivity to the rate of reproduction. Both measures varied by several fold as the temperature moved from 12C to 26C. As an interesting side-note, we found that the temperature of the agar was often several degrees lower than the thermostat temperature of the incubator or of enclosed flasks of water within the incubator, probably because of evaporative cooling at higher temperatures. Because of this, we report agar temperatures measured directly with a digital thermocouple thermometer. Net reproductive rate (total offspring production) displayed a large plateau in optimal performance from roughly 15-22C. The intrinsic rate of increase peaked at higher temperatures with a narrower plateau from 22-24C. The observation that r peaks at higher temperatures than R o appears to be a general phenomenon in ectotherms and is probably caused by r 's sensitivity to the rate of offspring production, which is substantially higher at higher temperatures. Before reaching a plateau, there is a roughly 40% increase in the rate of offspring production for every degree increase in temperature. Worms raised at 23C produce offspring over three times faster than worms raised at 12C. If selection for rapid reproduction is as important as it appears to be in C. elegans , then the effects of temperature on r should generate strong selection on many other aspects of the thermal ecology of these nematodes.

Archer, H. et al. (2017) International Worm Meeting "High throughput assessment of natural variation in the resistance to starvation stress in C. elegans using microfluidics."

Blue, B., Phillips, P.C., Banse, S., Archer, H.

[International Worm Meeting, 2017]

Caenorhabditis elegans typically feeds on rotting fruit and plant material in a fluctuating natural habitat, a boom-and-bust lifestyle. Moreover, stage specific developmental responses to low food concentration suggest that starvation-like conditions are a regular occurrence. In order to assess variation in the C. elegans starvation response under precisely controlled conditions and simultaneously phenotype a large number of individuals with high precision, we have developed a microfluidic device that, when combined with image scanning technology, allows for high-throughput assessment at a temporal resolution not previously feasible. Under these conditions worms exhibit a markedly reduced adult lifespan with strain-dependent variation in starvation resistance, ranging from <72 hours to ~120 hours. Initial results contrasting wildtype individuals with lipase mutants suggest a significant role for processes of fat metabolism in mediating resistance to starvation stress. Application of this device to a well-defined polymorphic population of C. elegans allows for mapping of the relationship between natural variation in genotypes and differences in starvation resistance.

Banse, S.A. et al. (2017) International Worm Meeting "The Crucible: A microfluidic platform for stress assays."

Jarret, C.M., Blue, B.W., Robinson, K.J., Phillips, P.C., Banse, S.A.

[International Worm Meeting, 2017]

In a world of uncertainty, an organism's ability to mount a response to stress is fundamental to its interaction with the environment. Experimental exploration of that interaction benefits greatly from tight environmental control, as well as the ability to minimize the behavioral response of fleeing stress. We present a microfluidic platform that enables tight environmental control, and high-throughput image acquisition while confining single animals to individual arenas that allow a broad range of behavioral activities. The platform is easily scalable, with two 50 arena arrays per chip, and an imaging capacity of 600 animals per imaging scanner. We present validation of the stress platform using osmotic stress, oxidative stress, and starvation, although the system should be adaptable to many stress paradigms.

Banse, S.A. et al. (2017) International Worm Meeting "Fine scale electrophysiological analysis of pharyngeal aging and a transition-state model of activity."

Lockery, S.R., Banse, S.A., Weeks, J.C., Willis, J.H., Phillips, P.C., Robinson, K.J., Blue, B.W.

[International Worm Meeting, 2017]

Because age-associated loss of neuromuscular function severely impacts quality of life, research models capable of identifying the etiology of lost function are particularly valuable. The C. elegans pharynx is a neuromuscular model whose visually observed age-dependent decline in contraction frequency is used as an experimental health marker. Although informative about overall health state, contraction frequency alone provides little information about the underlying cause for lost function. Fine scale analysis of electropharyngeograms recorded from aging animals provides an alternative assay more amenable to mapping age-related changes to the underlying neuromuscular circuit. We find that decreased pump frequency is associated with bursts of activity punctuated by increased frequency of pauses. Interestingly, although inter-pump interval during an activity bout changes with age, pause duration remains relatively unchanged. This suggests a decreased frequency or efficacy of cholinergic signaling during an activity bout. This reduced efficacy in neuronal modulation of pumping contrasts with M3 related glutamatergic signaling efficacy which holds longer into adulthood. This difference is visible both in the duration of pumps, as well as in average waveform shapes associated with the beginning and end of a pump. We present an activity-state based model to explain the observed decreased pump frequency. We argue that this new approach is an information dense measure that holds promise in mapping age-dependent changes to the functional subunits of pharyngeal activity.

Meneely, Philip M. et al. (2009) International Worm Meeting "HIM-8 interacts with SUN-1."

Daou, Maral, Thompson, Scott, Meneely, Philip M., Burch, Lauren

[International Worm Meeting, 2009]

During homolog recognition and pairing in meiosis, pairing centers at or near one end of each meiotic chromosome are localized to the nuclear envelope. SUN-1 is an evolutionarily conserved protein located in the inner nuclear membrane that appears to be directly involved in mediating telomere attachment in mice and yeast; in worms, a direct interaction between pairing center proteins and SUN-1 at the nuclear envelope has not yet been proved (1, 2). In sun-1 null mutants, none of the meiotic chromosomes are paired but the pairing centers are localized to the nuclear envelope (2). HIM-8 is a C2H2 zinc finger protein that binds specifically to the pairing center on the X chromosome and is necessary for the pairing of the X chromosome and for synapsis (3). When a zinc finger is mutated in him-8, such as in e1489 or the tm611 deletion, the protein does not bind to the X pairing center, the X chromosomes do not pair, but the chromosomes are still located at the nuclear envelope. Thus, pairing and nuclear envelope attachment may be separable activities for the pairing center proteins. The autosomes are paired normally in him-8 mutants. The three ZIM proteins, which are highly related to HIM-8 in sequence, mediate pairing of the autosomes (4). We find that SUN-1 and HIM-8 physically associate with each other in a yeast two-hybrid assay. Preliminary evidence suggests a similar physical interaction occurs between SUN-1 and at least one of the ZIM proteins. This suggests a model whereby meiotic chromosomes are tethered to the nuclear envelope for pairing via a direct interaction between SUN-1 in the inner membrane and HIM-8 or one of the ZIM proteins on each specific chromosome. 1.Penkner et al. 2007. Develop. Cell 12: 873-885 2.Fridkin, et al., 2009. Cell Mol. Life Sci. 66: (in press) 3.Phillips et al. 2005 Cell 123: 1051-1063 4.Phillips and Dernburg 2006 Develop. Cell 11: 817-829.

Yuriko Harigaya et al. (2007) International Worm Meeting "Assessing the ability of C.elegans spermatocytes to segregate achiasmate chromosomes."

Yuriko Harigaya, Anne Villeneuve

[International Worm Meeting, 2007]

Accurate segregation of homologous chromosomes at meiosis I is an essential step in sexual reproduction. This is usually achieved by a mechanism that relies on chiasmata, which are recombination-based linkages that temporarily connect homologs and allow them to orient toward opposite spindle pole. However, certain meiotic systems, such as those in Drosophila females and males, possess the ability to segregate homologs without chiasmata (achiasmate chromosome segregation) (Hawley et al. 1993). It has been proposed that this type of mechanism may operate in humans as a back up system in case of recombination failure (Koehler & Hassold 1998). Indirect genetic evidence has suggested that C.elegans spermatocytes may also have mechanisms that allow segregation of (at least) a single pair of achiasmate chromosomes (Albertson et al. 1997, for review). We have set out to revisit this issue, taking advantage of recent progress in the field of C. elegans meiosis. Phillips & Dernburg (2006) have identified a family of C2H2 zinc-finger proteins (ZIMs), which play a pivotal role in meiotic pairing and recombination. They have demonstrated that each of the three ZIM proteins act on specific autosome pairs in oocyte meiosis, that is, the zim mutations provide situations where one or two specific homologous pair(s) is/are consistently achiasmate. Results from the following analyses will be presented: 1) investigation of pairing and synapsis states in zim male germlines to confirm that ZIMs function similarly in spermatocyte meiosis; 2) testing cross progeny from the zim males to investigate whether achiasmate chromosome pairs, if any, are successfully inherited during meiosis in zim mutant spermatocytes; and 3) cytological observation of chromosomal placement at metaphase I and anaphase I in mutant spermatocytes using FISH analysis, to gain insights into the mechanistic bases for the chromosome inheritance pattern. Albertson et al. (1997) in C. ELEGANS II:p47; Hawley et al. (1993) Dev Genet 13:440; Koehler & Hassold (1998) Annu Hum Genet 62:467; Phillips & Dernburg (2006) Dev Cell 11:817.

Wynne, David et al. (2009) International Worm Meeting "Motor-Driven Motion Associated with Meiotic Chromosome Pairing."

Carlton, Pete, Wynne, David, Dernburg, Abby

[International Worm Meeting, 2009]

Accurate segregation of homologous chromosomes during meiosis is necessary for the faithful transmission of the genome from parent to progeny. To segregate properly, homologs must first undergo pairing, synapsis, and recombination. The mechanism of homologous chromosome pairing remains a major mystery of meiosis. In C. elegans, as in many other eukaryotes, pairing is accompanied by a global rearrangement of chromosomes. We have found that this rearrangement is driven through the association of special chromosome regions known as Pairing Centers (PC) with nuclear envelope proteins and cytoskeletal motors (Phillips et al. 2005, Sato A. unpublished). Using fluorescent markers for nuclear envelope attachment sites and Pairing Centers, we are analyzing chromosome dynamics through real-time imaging and quantitative motion tracking. Our results reveal a dramatic increase in chromosome motion at the onset of chromosome pairing that persists after homologous loci are paired. We show that this increased mobility is dependent on the conserved cell cycle checkpoint kinase, CHK-2, and that motion slows following knockdown of cytoplasmic dynein. Our data supports a model in which homolog pairing is promoted by a small number of fast, motor-driven movements that augment the smaller, Brownian motions seen prior to meiosis. The observation that fast motions persist well after pairing is completed suggests additional roles in chromosome synapsis or recombination.

Philipp Christoph Janiesch et al. (2008) European Worm Meeting "The Ubiquitin-Selective Chaperone CDC-48 Links Myosin Assembly in C. elegans to Human Myopathy"

Hanns Lochmuller, Thorsten Hoppe,, Philipp Christoph Janiesch, Giuseppe Cassata, Roja Barikbin, Johnny Kim, Sabine Krause, Julien Mouysset

[European Worm Meeting, 2008]

Protein degradation in eukaryotes often requires the ubiquitin-selective. chaperone p97 for substrate recruitment and ubiquitin chain assembly.. However, the physiological relevance of p97 and its role in developmental. processes are still unclear. Here, we discovered an unanticipated function. of CDC-48/p97 in myosin assembly and myofibril organization both in C.. elegans and humans. The developmentally regulated assembly of a CDC-48/UFD-. 2/CHN-1 complex links turnover of the myosin-directed chaperone UNC-45 to. functional muscle formation. Our data suggest a similarly conserved pathway. regulating myosin assembly in man. Remarkably, mutations in human p97,. known to cause hereditary inclusion body myopathy, abrogate UNC-45. degradation and result in severely disorganized myofibrils, detrimental. towards sarcomeric function. These results identify a key role of CDC-. 48/p97 in the process of myofiber differentiation and maintenance, which is. abolished during pathological conditions leading to protein aggregation and. inclusion body formation in human skeletal muscle. To our knowledge this is. the first study implicating functional myosin assembly and maintenance in. the pathophysiology of human inclusion body myopathy associated with Paget. disease of bone and frontotemporal dementia (IBMPFD).. The ubiquitin-selective chaperone CDC-48/p97 links myosin assembly to human. myopathy. Janiesch P.C., Kim J., Mouysset J., Barikbin R., Lochmuller H.,. Cassata G., Krause S., and Hoppe T. (2007). Nat. Cell Biol. 9, 379-390.. Multiubiquitylation by E4 enzymes: one size doesn''t fit all.. Hoppe T. (2005). Trends Biochem. Sci. 30, 183-187.

David Wynne et al. (2008) Development & Evolution Meeting "Real-Time Imaging of Meiotic Chromosome Dynamics"

Pete Carlton, David Wynne, Abby Dernburg

[Development & Evolution Meeting, 2008]

Accurate segregation of chromosomes during meiosis is necessary for the faithful transmission of the genome from parent to progeny. Conversely, meiotic missegregation leads to impaired viability and fertility, as well as aneuploid progeny that can have dramatic developmental defects, such as Down syndrome in humans. To segregate properly, homologs must first undergo pairing, synapsis, and recombination. Despite its central role in meiosis, pairing of homologous chromosomes remains poorly understood. In C. elegans, as in many other eukaryotes, this process is accompanied by a global rearrangement of chromosomes. We have found that this rearrangement is driven through the association of special chromosome regions, known as Homolog Recognition Regions or Pairing Centers, with the nuclear envelope. In other organisms, telomeres establish similar attachments to the NE during "bouquet stage" of meiosis, but the function of this configuration has not been established in any system. Chromosomes are not merely tethered at the nuclear envelope, but instead associate with specific nuclear envelope proteins and cytoskeletal motors (Phillips et al. 2005, Sato A. unpublished). Using ZYG-12:GFP (Malone et al. 2003) as a marker for the chromosome attachment sites, we have observed that they undergo rapid movements throughout the pairing process. Disruption of cytoplasmic dynein causes a dramatic decrease in patch motion, supporting a model in which pairing dynamics are driven by cytoskeletal components outside the nucleus. These data suggest that meiotic chromosome pairing is an active process rather than being purely a result of a diffusion-based mechanism.