Pu S et al. (2010) PLoS One "Expanding the landscape of chromatin modification (CM)-related functional domains and genes in human."

Pu S, On T, Zhang Z, Vlasblom J, Xiong X, Emili A, Turinsky AL, Wodak SJ, Greenblatt J, Parkinson J

[PLoS One, 2010]

Chromatin modification (CM) plays a key role in regulating transcription, DNA replication, repair and recombination. However, our knowledge of these processes in humans remains very limited. Here we use computational approaches to study proteins and functional domains involved in CM in humans. We analyze the abundance and the pair-wise domain-domain co-occurrences of 25 well-documented CM domains in 5 model organisms: yeast, worm, fly, mouse and human. Results show that domains involved in histone methylation, DNA methylation, and histone variants are remarkably expanded in metazoan, reflecting the increased demand for cell type-specific gene regulation. We find that CM domains tend to co-occur with a limited number of partner domains and are hence not promiscuous. This property is exploited to identify 47 potentially novel CM domains, including 24 DNA-binding domains, whose role in CM has received little attention so far. Lastly, we use a consensus Machine Learning approach to predict 379 novel CM genes (coding for 329 proteins) in humans based on domain compositions. Several of these predictions are supported by very recent experimental studies and others are slated for experimental verification. Identification of novel CM genes and domains in humans will aid our understanding of fundamental epigenetic processes that are important for stem cell differentiation and cancer biology. Information on all the candidate CM domains and genes reported here is publicly available.

Rincon S et al. (2019) J Infect Dis "Disrupting Membrane Adaptation Restores In Vivo Efficacy of Antibiotics Against Multidrug-Resistant Enterococci and Potentiates Killing by Human Neutrophils."

Cruz MR, Dinh AQ, Reyes J, Garsin DA, Arias CA, Khan A, Tran TT, Panesso D, Singh KV, Rincon S, Miller WR, Shamoo Y, Diaz L, Rios R

[J Infect Dis, 2019]

Daptomycin resistance in enterococci is often mediated by the LiaFSR system that orchestrates the cell membrane (CM) stress response. Activation of LiaFSR through the response regulator LiaR generates major changes in CM function and architecture (membrane adaptive response), permitting the organism to survive the antibiotic attack. Here, using a laboratory strain of Enterococcus faecalis, we developed a novel Caenorhabditis elegans model of daptomycin therapy and showed that disrupting LiaR-mediated CM adaptation restores the in vivo activity of daptomycin. The LiaR effect was also seen in a clinical strain of DAP-resistant E. faecium using a murine model of peritonitis. Furthermore, alteration of the CM response increased the ability of human-PMNs to readily clear both E. faecalis and MDR-E. faecium. Our results provide proof-of-concept that targeting the CM adaptive response restore the in vivo activity of antibiotics, prevent resistance and enhance the ability of the innate immune system to kill infecting bacteria.

White GE et al. (1993) International C. elegans Meeting "Synthesis of the major body wall muscle myosin in C. elegans is regulated by thick filament organization."

Schachat FH, Petry CM, White GE

[International C. elegans Meeting, 1993]

White GE et al. (2003) J Exp Biol "The pathway of myofibrillogenesis determines the interrelationship between myosin and paramyosin synthesis in Caenorhabditis elegans."

Schachat F, White GE, Petry CM

[J Exp Biol, 2003]

Examination of null mutants in myosin B and paramyosin yields insights into the complex mechanisms that regulate expression of the three major components of Caenorhabditis elegans body-wall muscle thick filaments myosin A, myosin B and paramyosin. In the absence of myosin B, paramyosin accumulation is reduced, although neither its synthesis nor that of myosin A is affected. This implies that the interaction of myosin B with paramyosin inhibits paramyosin degradation. By contrast, the absence of paramyosin results in reduced synthesis and accumulation of myosin B but has no effect on myosin A synthesis. The non-reciprocal effects of the null mutants on turnover and synthesis are best understood as an epigenetic phenomenon that reflects the pathway of thick filament assembly. The synthesis of myosin A and paramyosin, which are involved in the initial steps of thick filament formation, is independent of myosin B; however, a properly assembled paramyosin-containing thick filament core is essential for efficient synthesis of myosin B.

White GE et al. (1991) International C. elegans Meeting "BIOCHEMICAL AND MOLECULAR ANALYSIS OF THICK AND THIN FILAMENT PROTEIN EXPRESSION IN THE unc-78 MUTATION OF CAENORHABDITIS ELEGANS."

Petry CM, White GE, Schachat FH

[International C. elegans Meeting, 1991]

We are studying the effect of the unc-78 gene of C. elegans on muscle contractile protein expression. As described by Waterston, et al., (Dev. Bio. 77: 271,1980), the phenotype of unc-78 mutants is disruption of the myofilament lattice accompanied by accumulation of thin filaments at the periphery of body wall muscle cells of adult animals. Our goal is to understand whether this disruption results from alterations in transcription, posttranscriptional events, or assembly of muscle proteins. Our experimental approach has been to determine: l) protein accumulation profiles from myofibrillar preparations, 2) rates of protein synthesis from short term labelling of L3/L4 staged animals, and 3) relative mRNA levels for the contractile genes. Using quantitative densitometry, we have determined the levels of expression of four major components of the myofilarnent lattice: actin, myosin, tropomyosin, and paramyosin. Comparison of myofibrillar preps from CB1217 (the reference allele for unc-78) and the wild type, N2, by SDS-PAGE shows that the levels of both myosin and paramyosin are reduced by approximately 25% in CB1217. Analysis of the myosin heavy chains using the Neville gel system reveals that the ratio of the two myosins of the body wall muscle, myosin A and myosin B, is not significantly different from the wild type. Myofibrillar profiles of the unc-78 suppressor strain RW2337 (sup-13 (kindly provided by R.H. Waterston)), reveals that myosin and paramyosin expression is significantly increased in this suppressor strain. This implies that the disruption of the myofilarnent lattice that occurs in unc-78 mutants results from under-expression of thick filament proteins. Initial studies on the rates of protein synthesis for the myofibrillar proteins indicate a slight difference between the wild type and unc-78 animals. Studies on sup-13 animals indicate that it suppresses unc-78 mutations by increasing the rate of synthesis of myosin and paramyosin. Preliminary studies of message levels for these same four contractile proteins reveals differences between unc-78, sup- 13, and the wild type.

On T et al. (2010) Proteins "The evolutionary landscape of the chromatin modification machinery reveals lineage specific gains, expansions, and losses."

On T, Emili A, Turinsky A, Zhang Z, Xiong X, Greenblatt J, Gong Y, Wodak SJ, Parkinson J, Pu S

[Proteins, 2010]

Model organisms such as yeast, fly, and worm have played a defining role in the study of many biological systems. A significant challenge remains in translating this information to humans. Of critical importance is the ability to differentiate those components where knowledge of function and interactions may be reliably inferred from those that represent lineage-specific innovations. To address this challenge, we use chromatin modification (CM) as a model system for exploring the evolutionary properties of their components in the context of their known functions and interactions. Collating previously identified components of CM from yeast, worm, fly, and human, we identified a "core" set of 50 CM genes displaying consistent orthologous relationships that likely retain their interactions and functions across taxa. In addition, we catalog many components that demonstrate lineage specific expansions and losses, highlighting much duplication within vertebrates that may reflect an expanded repertoire of regulatory mechanisms. Placed in the context of a high-quality protein-protein interaction network, we find, contrary to existing views of evolutionary modularity, that CM complex components display a mosaic of evolutionary histories: a core set of highly conserved genes, together with sets displaying lineage specific innovations. Although focused on CM, this study provides a template for differentiating those genes which are likely to retain their functions and interactions across species. As such, in addition to informing on the evolution of CM as a system, this study provides a set of comparative genomic approaches that can be generally applied to any biological systems.

Khan A et al. (2019) Proc Natl Acad Sci U S A "Antimicrobial sensing coupled with cell membrane remodeling mediates antibiotic resistance and virulence in Enterococcus faecalis."

Nguyen AH, Reyes J, Narechania A, Tran TT, Shamoo Y, Siegel SD, Rios R, Ton-That H, Cruz MR, Singh KV, Arias CA, Garsin DA, Khan A, Rincon S, Miller WR, Pemberton O, Diaz L, Latorre M, Davlieva M, Planet PJ, Panesso D

[Proc Natl Acad Sci U S A, 2019]

Bacteria have developed several evolutionary strategies to protect their cell membranes (CMs) from the attack of antibiotics and antimicrobial peptides (AMPs) produced by the innate immune system, including remodeling of phospholipid content and localization. Multidrug-resistant <i>Enterococcus faecalis,</i> an opportunistic human pathogen, evolves resistance to the lipopeptide daptomycin and AMPs by diverting the antibiotic away from critical septal targets using CM anionic phospholipid redistribution. The LiaFSR stress response system regulates this CM remodeling via the LiaR response regulator by a previously unknown mechanism. Here, we characterize a LiaR-regulated protein, LiaX, that senses daptomycin or AMPs and triggers protective CM remodeling. LiaX is surface exposed, and in daptomycin-resistant clinical strains, both LiaX and the N-terminal domain alone are released into the extracellular milieu. The N-terminal domain of LiaX binds daptomycin and AMPs (such as human LL-37) and functions as an extracellular sentinel that activates the cell envelope stress response. The C-terminal domain of LiaX plays a role in inhibiting the LiaFSR system, and when this domain is absent, it leads to activation of anionic phospholipid redistribution. Strains that exhibit LiaX-mediated CM remodeling and AMP resistance show enhanced virulence in the <i>Caenorhabditis elegans</i> model, an effect that is abolished in animals lacking an innate immune pathway crucial for producing AMPs. In conclusion, we report a mechanism of antibiotic and AMP resistance that couples bacterial stress sensing to major changes in CM architecture, ultimately also affecting host-pathogen interactions.

Pang S et al. (2010) Lab Chip "Implementation of a color-capable optofluidic microscope on a RGB CMOS color sensor chip substrate."

Wang YM, Yang C, DeModena J, Cui X, Sternberg P, Pang S

[Lab Chip, 2010]

We report the implementation of a color-capable on-chip lensless microscope system, termed color optofluidic microscope (color OFM), and demonstrate imaging of double stained Caenorhabditis elegans with lacZ gene expression at a light intensity about 10 mW/cm(2).

Chris Gabel et al. (2004) East Coast Worm Meeting "Quantification of electrotaxis"

Dmitri Pavlichin, Chris Gabel, Aravi Samuel, Albert Kao

[East Coast Worm Meeting, 2004]

In an electric field, the worm has preferred trajectories for forward crawling movement that depend on the direction and magnitude of the electric field. In weak fields (<5 V/cm), worms tend to crawl towards either the positive or negative pole. In moderate fields, ~5 V/cm, worms crawl straight towards the negative pole. In stronger fields, >5 V/cm, worms crawl towards the negative pole in trajectories at an angle to the direction of the field. The angle of approach towards the negative pole increases with field strength, rising from 0 degrees (parallel to the field lines) towards 90 degrees (perpendicular to the field lines). We have quantified these electrotactic movements by tracking wild-type and mutant worms crawling over agar surfaces containing different types and concentrations of ions while responding to electric fields varied in amplitude, frequency, and direction.

Mendoza, S. et al. (2017) International Worm Meeting "W-TEM: A microscope to study C. elegans behavior under external stimulations."

Lam, B., Mendoza, S., D'Orazio, E., Sherry, T., Madruga, B., Mai, P., Arisaka, K., Jiang, K.

[International Worm Meeting, 2017]

The ability to monitor neural activity in freely behaving animals is important in determining neural mechanisms responsible for behavior. Real time worm tracking with neuronal observation has been used to study worm behavior in freely moving C. elegans. However, few setups have attempted to use worm tracking in conjunction with behavioral stimulations. The difficulty being that most behavioral stimulations used to study C. elegans such as electrotaxis and thermotaxis are difficult to incorporate into existing worm tracking platforms. We present a novel microscope platform, W-TEM ( Worm Tracking Epifluorescence Microscope), to bridge this gap in our understanding of worm behavior. The microscope is a standard wide field epi-fluorescence microscope placed on top of a stable platform to move the microscope as it tracks along the sample plane, accommodating most C. elegans behavioral stimulations. This microscope can incorporate a stimulation system that covers 20 cm x 20 cm of worm area movement, 50 cm x 50 cm of total size, and a height of up to 30 cm, which is larger than most other stimulation systems. It can take data at video rate 30 fps, for cameleon ratiometic imaging, as well as monitor whole worm behavior. In addition, our platform can be moved easily between different experiments, using a system of rails that is attached to the microscope platform. Using this system, we have been able to study both Electrotaxis and Thermotactic behaviors using the same system. In the case of Electrotaxis, we have found that high voltages corresponding to 8V/cm create a dampening effect of worm motion, possibly due to paralyzing the lower half of the worm body. We also conduct investigations into the AFD and AIY neurons during isothermal behavior and find a phase lag of 0.8 seconds between neural activity and head location during isothermal tracking.