Aguilar GR et al. (2024) MicroPubl Biol "A protocol to transform a fluorescent reporter from a nuclear to a cytoplasmic location."

Hobert O, Aguilar GR

[MicroPubl Biol, 2024]

To facilitate cell identification for expression pattern analysis in <i>C. elegans</i> , an SL2::GFP::H2B fluorescent reporter cassette has become a popular and widely used choice to generate nuclear localized reporter alleles by CRISPR/Cas9 genome engineering. When added at the 3' end of a locus of interest, this cassette concentrates GFP into the nucleus and permits the identification of expressing cells, for example with the help of the NeuroPAL tool. However, there are instances in which it is desirable to visualize the complete morphology of a cell that expresses an SL2::GFP::H2B reporter cassette. We describe here a CRISPR/Cas9-engineering strategy to transform an endogenous SL2::GFP::H2B tag into a cytosolic tag by insertion of the self-cleaving T2A tag in between GFP and H2B.

Aguilar G et al. (2019) Antibodies (Basel) "Using Nanobodies to Study Protein Function in Developing Organisms."

Matsuda S, Aguilar G, Vigano MA, Affolter M

[Antibodies (Basel), 2019]

Polyclonal and monoclonal antibodies have been invaluable tools to study proteins over the past decades. While indispensable for most biological studies including developmental biology, antibodies have been used mostly in fixed tissues or as binding reagents in the extracellular milieu. For functional studies and for clinical applications, antibodies have been functionalized by covalently fusing them to heterologous partners (i.e., chemicals, proteins or other moieties). Such functionalized antibodies have been less widely used in developmental biology studies. In the past few years, the discovery and application of small functional binding fragments derived from single-chain antibodies, so-called nanobodies, has resulted in novel approaches to study proteins during the development of multicellular animals in vivo. Expression of functionalized nanobody fusions from integrated transgenes allows manipulating proteins of interest in the extracellular and the intracellular milieu in a tissue- and time-dependent manner in an unprecedented manner. Here, we describe how nanobodies have been used in the field of developmental biology and look into the future to imagine how else nanobody-based reagents could be further developed to study the proteome in living organisms.

Aguilar PS et al. (2013) Trends Genet "Genetic basis of cell-cell fusion mechanisms."

Podbilewicz B, Helming L, Fleissner A, Aguilar PS, Inoue N, Wang H, Baylies MK, Wong M

[Trends Genet, 2013]

Cell-cell fusion in sexually reproducing organisms is a mechanism to merge gamete genomes and, in multicellular organisms, it is a strategy to sculpt organs, such as muscle, bone, and placenta. Moreover, this mechanism has been implicated in pathological conditions, such as infection and cancer. Studies of genetic model organisms have uncovered a unifying principle: cell fusion is a genetically programmed process. This process can be divided in three stages: competence (cell induction and differentiation); commitment (cell determination, migration, and adhesion); and cell fusion (membrane merging and cytoplasmic mixing). Recent work has led to the discovery of fusogens, which are cell fusion proteins that are necessary and sufficient to fuse cell membranes. Two unrelated families of fusogens have been discovered, one in mouse placenta and one in Caenorhabditis elegans (syncytins and F proteins, respectively). Current research aims to identify new fusogens and determine the mechanisms by which they merge membranes.

Camacho-Aguilar E et al. (2021) PLoS Comput Biol "Quantifying cell transitions in C. elegans with data-fitted landscape models."

Rand DA, Warmflash A, Camacho-Aguilar E

[PLoS Comput Biol, 2021]

Increasing interest has emerged in new mathematical approaches that simplify the study of complex differentiation processes by formalizing Waddington's landscape metaphor. However, a rational method to build these landscape models remains an open problem. Here we study vulval development in C. elegans by developing a framework based on Catastrophe Theory (CT) and approximate Bayesian computation (ABC) to build data-fitted landscape models. We first identify the candidate qualitative landscapes, and then use CT to build the simplest model consistent with the data, which we quantitatively fit using ABC. The resulting model suggests that the underlying mechanism is a quantifiable two-step decision controlled by EGF and Notch-Delta signals, where a non-vulval/vulval decision is followed by a bistable transition to the two vulval states. This new model fits a broad set of data and makes several novel predictions.

Aguilar-Portillo, Georgina et al. (2013) International Worm Meeting "The Pristionchus pacificus obi-3 mutant lacks attraction to beetle host pheromone and shows increased turning frequency."

Muthumala, Neomal, Hong, Ray, Escobedo, Jimmy, Aguilar-Portillo, Georgina

[International Worm Meeting, 2013]

Understanding chemosensation and neurophysiology from varying nematode species is crucial for investigating gene function evolution. The soil-dwelling, free-living C. elegans has elucidated key neurons and proteins required for chemosensation of volatile odors, pheromones, and food. A more recently discovered nematode species, Pristionchus pacificus has been found to associate with specific beetle hosts, including the oriental beetle found in Japan and northeastern United States, likely through attraction to its sex pheromone, ZTDO. To determine the genes responsible for insect pheromone sensing, two chemosensory mutants were identified in a genetic screen for lack of attraction towards ZTDO after cGMP treatment and were named obi-1 and obi-3 (oriental beetle pheromone insensitive). This study focuses on obi-3, which also has a short body length and locomotion phenotype. Wild-type P. pacificus reverses 3x more frequently as C. elegans and reversal rates are known to affect approach behavior in chemotaxis. Obi-3's higher reversal rate was quantified by counting omega turns on and off food. An omega turn is defined by a worm turning its head 180 deg , touching the tip of its tail, and proceeding in a forward motion in a new direction. Interestingly, C. elegans egl-30 mutants show a similar higher reversal rate. Defects in EGL-30 affects the Gqa pathway that mediates the release of acetylcholine from motor neurons in order for normal muscle movement to occur. We also found that the associated coiling behavior is further accentuated in liquid medium. Wild-type shows a thrashing motion whereas obi-3 coils into a superimposed circle. This coiling-swimming phenotype was utilized for positional mapping in obi-3 x Japan recombinant inbred lines and rough mapping the obi-3 mutation to 38 cM interval of Chromosome I using SSLP markers. The long-term goal is to determine if obi-3 and obi-1 constitute components of an unknown signaling pathway, part of the conserved cGMP-dependent G protein signaling pathway, or TRPV channels lipid signaling pathway involved in sensing insect hosts. [This study was supported by NIH SCORE SC2GM089602 and NSF HRD-1139803 CSU-LSAMP Bridge to the Doctorate].

Haji Adineh, S et al. (2019) International Worm Meeting "The tempo of paternal mitochondrial transmission in C. briggsae hybrids."

Jorgensen, C, Chahal, G, Haji Adineh, S, Ortega, B, Aguilar, A, Ross, J

[International Worm Meeting, 2019]

Although mitochondria are usually maternally inherited, substantial evidence from multiple taxa reveals that hybridization often promotes transmission of sperm-borne mitochondrial into the fertilized oocyte. Given prior reports of such paternal "leakage" of mitochondria in intra-species C. briggsae crosses, we conducted new experiments to refine our understanding of the genetic basis and tempo of paternal mitochondrial transmission (PMT). Analysis of paternal mitochondrial haplotypes in replicate hybrid lines revealed persistent but transient heteroplasmy that never persisted beyond ten generations. Thus, PMT appears to occur regularly following hybridization, but heteroplasmy is usually resolved in favor of the maternal mitotype. Additionally, no paternal mitotypes were detected among a sample of existing C. briggsae advanced-intercross recombinant inbred lines (AI-RIL), which could suggest that using recombination to disrupt a potentially complex genetic mechanism that regularly acts to prevent paternal transmission does not result in sustained homoplasmy of the paternal mitotype. However, crosses involving AI-RIL pseudofemales do occasionally result in PMT, further implicating the role of a hybrid maternal nuclear genome in facilitating PMT. Current efforts in C. elegans are testing the hypothesis that PMT is dependent on somatic sex, such that hermaphrodites do not perform paternal mitochondrial elimination during self-fertilization but do eliminate non-self sperm mitochondria when mated with a male.

Quartey MO et al. (2021) Sci Rep "The A(1-38) peptide is a negative regulator of the A(1-42) peptide implicated in Alzheimer disease progression."

Pennington PR, Heistad RM, Nyarko JNK, Barnes JR, Bolanos MAC, Parsons MP, Knudsen KJ, De Carvalho CE, Leary SC, Mousseau DD, Buttigieg J, Maley JM, Quartey MO

[Sci Rep, 2021]

The pool of -Amyloid (A) length variants detected in preclinical and clinical Alzheimer disease (AD) samples suggests a diversity of roles for A peptides. We examined how a naturally occurring variant, e.g. A(1-38), interacts with the AD-related variant, A(1-42), and the predominant physiological variant, A(1-40). Atomic force microscopy, Thioflavin T fluorescence, circular dichroism, dynamic light scattering, and surface plasmon resonance reveal that A(1-38) interacts differently with A(1-40) and A(1-42) and, in general, A(1-38) interferes with the conversion of A(1-42) to a -sheet-rich aggregate. Functionally, A(1-38) reverses the negative impact of A(1-42) on long-term potentiation in acute hippocampal slices and on membrane conductance in primary neurons, and mitigates an A(1-42) phenotype in Caenorhabditis elegans. A(1-38) also reverses any loss of MTT conversion induced by A(1-40) and A(1-42) in HT-22 hippocampal neurons and APOE 4-positive human fibroblasts, although the combination of A(1-38) and A(1-42) inhibits MTT conversion in APOE 4-negative fibroblasts. A greater ratio of soluble A(1-42)/A(1-38) [and A(1-42)/A(1-40)] in autopsied brain extracts correlates with an earlier age-at-death in males (but not females) with a diagnosis of AD. These results suggest that A(1-38) is capable of physically counteracting, potentially in a sex-dependent manner, the neuropathological effects of the AD-relevant A(1-42).

Danielle Thierry-Mieg et al. (2003) Worm Breeder's Gazette "www.wormgenes.org , a new gene centered database"

Vahan Simonyan, Michel Potdevin, Mark Sienkiewicz, Yuji KOHARA, Jean Thierry-Mieg, Danielle Thierry-Mieg, Tadasu SHIN-I

[Worm Breeder's Gazette, 2003]

Wormgenes is a new resource for C.elegans offering a detailed summary about each gene and a powerful query system.

Tangrodchanapong T et al. (2020) Front Pharmacol "Frondoside A Attenuates Amyloid- Proteotoxicity in Transgenic Caenorhabditis elegans by Suppressing Its Formation."

Tangrodchanapong T, Sobhon P, Meemon K

[Front Pharmacol, 2020]

Oligomeric assembly of Amyloid- (A) is the main toxic species that contribute to early cognitive impairment in Alzheimer's patients. Therefore, drugs that reduce the formation of A oligomers could halt the disease progression. In this study, by using transgenic <i>Caenorhabditis elegans</i> model of Alzheimer's disease, we investigated the effects of frondoside A, a well-known sea cucumber <i>Cucumaria frondosa</i> saponin with anti-cancer activity, on A aggregation and proteotoxicity. The results showed that frondoside A at a low concentration of 1 M significantly delayed the worm paralysis caused by A aggregation as compared with control group. In addition, the number of A plaque deposits in transgenic worm tissues was significantly decreased. Frondoside A was more effective in these activities than ginsenoside-Rg3, a comparable ginseng saponin. Immunoblot analysis revealed that the level of small oligomers as well as various high molecular weights of A species in the transgenic <i>C. elegans</i> were significantly reduced upon treatment with frondoside A, whereas the level of A monomers was not altered. This suggested that frondoside A may primarily reduce the level of small oligomeric forms, the most toxic species of A. Frondoside A also protected the worms from oxidative stress and rescued chemotaxis dysfunction in a transgenic strain whose neurons express A. Taken together, these data suggested that low dose of frondoside A could protect against A-induced toxicity by primarily suppressing the formation of A oligomers. Thus, the molecular mechanism of how frondoside A exerts its anti-A aggregation should be studied and elucidated in the future.

Hodgkin JA (1998) International Journal of Developmental Biology "Seven types of pleiotropy."

Hodgkin JA

[International Journal of Developmental Biology, 1998]

Pleiotropy , a situation in which a single gene influences multiple phenotypic tra its, can arise in a variety of ways. This paper discusses possible underlying mechanisms and proposes a classification of the various phenomena involved.