Purice, Maria et al. (2021) International Worm Meeting "The molecular atlas of C. elegans glia across sex and age"

Wu, Feinan, Singhvi, Aakanksha, Sorrentino, Violet, Setty, Manu, Purice, Maria

[International Worm Meeting, 2021]

Interactions between neurons and glia are essential for all normal nervous system functions. C. elegans glia have similar types of function as mammalian glia and are now established as a powerful genetic model to study glial biology. The adult hermaphrodite nervous system contains 302 neurons and 56 glia, with at least some of the glia shown to profoundly impact associated neuron shape and associated animal behaviors. Adult males have an additional 36 glia and 89 neurons, many of which control male mating-related behaviors. A major limitation in studying C. elegans glia functions is the lack of cell-type specific markers for each glial subtype in the adult worm, and a global comparative assessment of glial functional and molecular heterogeneity. Further, each C. elegans glia is born of an invariant developmental lineage and makes invariant neuron-contact. This not only makes the nematode a powerful model to study glia-neuron interactions, but also suggests a unique advantage of this model to examine glia at single-cell and molecular detail. Addressing this gap, we present our single nuclear RNAseq (snRNAseq) studies on adult C. elegans glia across the entire nervous system. We have done these studies in three settings, (1) young adult hermaphrodites, (2) young adult males, and (3) aged hermaphrodites. These data will be compiled and shared as a searchable three-dimensional online atlas of adult glial gene expression as a community resource. Our datasets allow us to now create a gene expression and marker atlas of glia at single cell resolution. Our preliminary data reveal 30 clusters, hinting already at interesting biological insights into glial heterogeneity. Further, studies in our lab and others have found that multiple glial cues within a single glia, the amphid sheath (AMsh), regulate neuron morphology and sensory behaviors, with age-related effects. Therefore, in addition to describing glia-specific and glial subclass markers, we are currently validating our datasets functionally in similar assays, including neural aging. The aim of our studies is to expand our understanding of how different glia execute these regulatory neural functions across cell-type, sex and animal aging.

De Magalhaes Filho CD et al. (2018) Nat Commun "Visible light reduces C. elegans longevity."

Dillin A, Seah NE, Henriquez B, Lapierre LR, Evans RM, De Magalhaes Filho CD

[Nat Commun, 2018]

The transparent nematode Caenorhabditis elegans can sense UV and blue-violet light to alter behavior. Because high-dose UV and blue-violet light are not a common feature outside of the laboratory setting, we asked what role, if any, could low-intensity visible light play in C. elegans physiology and longevity. Here, we show that C. elegans lifespan is inversely correlated to the time worms were exposed to visible light. While circadian control, lite-1 and tax-2 do not contribute to the lifespan reduction, we demonstrate that visible light creates photooxidative stress along with a general unfolded-protein response that decreases the lifespan. Finally, we find that long-lived mutants are more resistant to light stress, as well as wild-type worms supplemented pharmacologically with antioxidants. This study reveals that transparent nematodes are sensitive to visible light radiation and highlights the need to standardize methods for controlling the unrecognized biased effect of light during lifespan studies in laboratory conditions.

Klass MR (1977) Mech Ageing Dev "Aging in the nematode Caenorhabditis elegans: major biological and environmental factors influencing life span."

Klass MR

[Mech Ageing Dev, 1977]

The free-living nematode Caenorhabditis elegans is an excellent experimental system for the study of aging. The present study identifies some of the major biological and environmental factors influencing life span as a prelude to more detailed genetic and biochemical analyses. Life span can be altered during any part of the life cycle by a change in either temperature or food concentration. Parental age and parental life span both have relatively small effects on progeny life span. The nematode accumulates fluorescent pigment resembling lipofuscin, and becomes less sensitive to ultra-violet radiation as it

Zhou XX et al. (2017) Science "Optical control of cell signaling by single-chain photoswitchable kinases."

Shen K, Li P, Fan LZ, Lin MZ, Zhou XX

[Science, 2017]

Protein kinases transduce signals to regulate a wide array of cellular functions in eukaryotes. A generalizable method for optical control of kinases would enable fine spatiotemporal interrogation or manipulation of these various functions. We report the design and application of single-chain cofactor-free kinases with photoswitchable activity. We engineered a dimeric protein, pdDronpa, that dissociates in cyan light and reassociates in violet light. Attaching two pdDronpa domains at rationally selected locations in the kinase domain, we created the photoswitchable kinases psRaf1, psMEK1, psMEK2, and psCDK5. Using these photoswitchable kinases, we established an all-optical cell-based assay for screening inhibitors, uncovered a direct and rapid inhibitory feedback loop from ERK to MEK1, and mediated developmental changes and synaptic vesicle transport in vivo using light.

Park HK et al. (2004) DNA Repair (Amst) "A DNA repair gene of Caenorhabditis elegans: a homolog of human XPF."

Park HK, Ahn B, Suh D, Koo HS, Hyun M

[DNA Repair (Amst), 2004]

The xeroderma pigmentosum complementation group F (XPF) protein is a structure-specific endonuclease in a complex with ERCC1 and is essential for nucleotide excision repair (NER). We report a single cDNA of Caenorhabditis elegans (C. elegans) encoding highly similar protein to human XPF and other XPF members. We propose to name the corresponding C. elegans gene xpf. Messenger RNA for C. elegans xpf is 5''-tagged with a SL2 splice leader, suggesting an operon-like expression for xpf. Using RNAi, we showed that loss of C. elegans xpf function caused hypersensitivity to ultra-violet (UV) irradiation, as observed in enhanced germ cell apoptosis and increased embryonic lethality. This study suggests that C. elegans xpf is conserved in evolution and plays a role in the repair of UV-damaged DNA in C. elegans.

Viszwapriya D et al. (2016) Pathog Dis "Betulin inhibits virulence and biofilm of Streptococcus pyogenes by suppressing ropB core regulon, sagA and dltA."

Balamurugan K, Prithika U, Subramenium GA, Pandian SK, Viszwapriya D

[Pathog Dis, 2016]

The present study demonstrates the antivirulence potential of betulin, an abundantly available triterpenoid against Streptococcus pyogenes, a multi-virulent and exclusive human pathogen. Crystal violet assay and microscopic examination revealed that betulin (100 g mL(-1)) exhibits surface independent antibiofilm activity and mitigates extracellular polymeric substance production. Betulin treatment enhanced the rate of auto-aggregation in liquid medium. Results of real-time PCR and biochemical assays demonstrated that, betulin suppresses the expression of ropB core regulon, sagA and dltA, which correspondingly affects SpeB production, hemolysis and cell surface hydrophobicity, for the observed impairment in virulence and biofilm formation. dltA down regulation also affected the production of M protein, making betulin treated cells more susceptible to phagocytosis. The non-toxic nature of betulin and its antivirulence potential against S. pyogenes was manifested in vivo in Caenorhabditis elegans This study reveals the prospective role of betulin as therapeutic agent for the prevention and treatment of streptococcal infections.

Seung-hwan Kim et al. (2005) International Worm Meeting "The kinetochore components HCP-1 and HCP-2 promote S phase progression when chromosomes are damaged during early embryogenesis in C. elegans."

W Matthew Michael, Seung-hwan Kim

[International Worm Meeting, 2005]

We performed a systematic screen using RNAi of 4149 genes on chromosome V to identify those genes whose loss-of-function by RNAi causes sensitivity to two DNA damaging agents, methyl methanesulfonate (MMS) and ultra violet (UV) light. These MMS- and UV-sensitive genes fell into several categories, such as cell cycle and checkpoint regulators, DNA repair genes, and meiotic genes. Among the genes recovered in this screen were hcp-1 and hcp-2, which have previously been shown to function in kinetochore assembly. Depletion of either hcp-1 or hcp-2 by RNAi caused two phenotypes that were dependent upon DNA damage. One, progression through S phase in the early cleavage cycles was significantly delayed. Two, nuclear morphology in early embryos was perturbed in that nuclei were enlarged and of abnormal shape. These findings link kinetochore assembly to S phase progression during the DNA damage response and suggest a feedback mechanism whereby DNA synthesis is slowed when kinetochore assembly is attenuated on damaged chromosomes.

Kaplan JB et al. (2012) J Antibiot (Tokyo) "Recombinant human DNase I decreases biofilm and increases antimicrobial susceptibility in staphylococci."

Kaplan JB, Cardona ST, Lovetri K, Madhyastha S, Jabbouri S, Izano EA, Sadovskaya I

[J Antibiot (Tokyo), 2012]

Extracellular DNA is an adhesive component of staphylococcal biofilms. The aim of this study was to evaluate the antibiofilm activity of recombinant human DNase I (rhDNase) against Staphylococcus aureus and Staphylococcus epidermidis. Using a 96-well microtiter plate crystal-violet binding assay, we found that biofilm formation by S. aureus was efficiently inhibited by rhDNase at 1-4gl, and preformed S. aureus biofilms were efficiently detached in 2min by rhDNase at 1mgl. Pretreatment of S. aureus biofilms for 10min with 10mgl rhDNase increased their sensitivity to biocide killing by 4-5 log units. rhDNase at 10mgl significantly inhibited biofilm formation by S. epidermidis in medium supplemented with sub-MICs of antibiotics. We also found that rhDNase significantly increased the survival of S. aureus-infected Caenorhabditis elegans nematodes treated with tobramycin compared with nematodes treated with tobramycin alone. We concluded that rhDNase exhibits potent antibiofilm and antimicrobial-sensitizing activities against S. aureus and S. epidermidis at clinically achievable concentrations. rhDNase, either alone or in combination with antimicrobial agents, may have applications in treating or preventing staphylococcal biofilm-related infections.

Jung HI et al. (2017) J Microbiol "Mutation of the cyclic di-GMP phosphodiesterase gene in Burkholderia lata SK875 attenuates virulence and enhances biofilm formation."

Kim YJ, Jung HI, Lee HS, Kim SK, Lee JK, Lee YJ

[J Microbiol, 2017]

Burkholderia sp. is a gram-negative bacterium that commonly exists in the environment, and can cause diseases in plants, animals, and humans. Here, a transposon mutant library of a Burkholderia lata isolate from a pig with swine respiratory disease in Korea was screened for strains showing attenuated virulence in Caenorhabditis elegans. One such mutant was obtained, and the Tn5 insertion junction was mapped to rpfR, a gene encoding a cyclic di-GMP phosphodiesterase that functions as a receptor. Mutation of rpfR caused a reduction in growth on CPG agar and swimming motility as well as a rough colony morphology on Congo red agar. TLC analysis showed reduced AHL secretion, which was in agreement with the results from plate-based and bioluminescence assays. The mutant strain produced significantly more biofilm detected by crystal violet staining than the parent strain. SEM of the mutant strain clearly showed that the overproduced biofilm contained a filamentous structure. These results suggest that the cyclic di-GMP phosphodiesterase RpfR plays an important role in quorum sensing modulation of the bacterial virulence and biofilm formation.

Fang H et al. (2018) Front Cell Infect Microbiol "BfvR, an AraC-Family Regulator, Controls Biofilm Formation and pH6 Antigen Production in Opposite Ways in Yersinia pestis Biovar Microtus."

Yang R, Fang H, Yan Y, Liu L, Zhou D, Yang H, Han Y, Ding X, Zhang Y

[Front Cell Infect Microbiol, 2018]

Biofilm formation is critical for blocking flea foregut and hence for transmission of <i>Y. pestis</i> by flea biting. In this study, we identified the regulatory role of the AraC-family transcriptional regulator BfvR (YPO1737 in strain CO92) in biofilm formation and virulence of <i>Yersinia pestis</i> biovar Microtus. Crystal violet staining, <i>Caenorhabditis elegans</i> biofilm assay, colony morphology assay, intracellular c-di-GMP concentration determination, and BALB/c mice challenge were employed to reveal that BfvR enhanced <i>Y. pestis</i> biofilm formation while repressed its virulence in mice. Further molecular biological assays demonstrated that BfvR directly stimulated the expression of <i>hmsHFRS, waaAE-coaD</i>, and <i>hmsCDE</i>, which, in turn, affected the production of exopolysaccharide, LPS, and c-di-GMP, respectively. In addition, BfvR directly and indirectly repressed <i>psaABC</i> and <i>psaEF</i> transcription, respectively. We concluded that the modulation of biofilm- and virulence-related genes by BfvR led to increased biofilm formation and reduced virulence of <i>Y. pestis</i> biovar Microtus.