Ceballos, Ainhoa et al. (2021) MicroPubl Biol

Ceballos, Ainhoa, Esse, Ruben, Grishok, Alla

[MicroPubl Biol, 2021]

The only representative of the MYC superfamily transcription factors in C. elegans, MML-1 (Myc and Mondo-like 1), was shown to promote extended lifespan in a variety of models and to regulate some aspects of C. elegans development. This previous research did not involve molecular characterization of MML-1. Here we use available mml-1 mutant alleles and other reagents to demonstrate that MML-1 is modified by O-GlcNAc, binds to promoters of some genes directly regulated by the DOT-1.1 histone methyltransferase complex, and has a role in promoting neuronal migration. Surprisingly, we found that the deletion allele mml-1(ok849), which was considered a null, produces an internally truncated protein resulting from an in-frame deletion. Localization of this truncated product to MML-1 target promoters was not impaired. The deleted region of MML-1 is proline-rich, and its function is poorly understood in mammalian homologs of MML-1. Based on our work and previously published data we conclude that the internal proline-rich region of MML-1 is dispensable for DNA binding but is biologically important.

Ceballos AV et al. (2018) Methods Enzymol "Methods and Strategies to Quantify Phase Separation of Disordered Proteins."

Elbaum-Garfinkle S, Ceballos AV, McDonald CJ

[Methods Enzymol, 2018]

Phase separation has emerged as a new paradigm currently revolutionizing our understanding of cell biology and intracellular organization. Disordered protein domains have recently been demonstrated as integral drivers of phase separation into condensed liquids with emergent material properties. Using in vitro model systems employing purified protein components is necessary to interrogate the molecular mechanisms underlying phase separation; however, these systems pose many experimental challenges. In this chapter we describe general strategies for purifying, handling, imaging, and characterizing the phase behavior of disordered proteins. We further outline methods for the purification of the model P granule protein LAF-1, the construction of phase diagrams, and the quantification of liquid droplet fusion or coalescence.

Ceballos, Ainhoa et al. (2015) International Worm Meeting "Transcriptional regulation of metabolic genes by promoter-localized O-GlcNAc."

Cecere, Germano, Ceballos, Ainhoa, Grishok, Alla

[International Worm Meeting, 2015]

The O-Linked-N-acetylglucosamine (O-GlcNAc) modification of proteins is one of the most abundant in eukaryotes. It is very sensitive to glucose levels and environmental conditions and therefore has the potential to lead environmentally-induced epigenetic changes. In C. elegans, prominent peaks of O-GlcNAc were identified close to transcription start sites (TSS) of many metabolic genes by ChIP/chip, although the identitiy of the O-GlcNAc modified protein is unknown (Love et al.,2010). We described a negative regulation of transcription of growth-related and metabolic genes by the conserved chromatin-binding factor ZFP-1 and its interacting partner the histone methyltransferase DOT-1.1 (Mansisidor et al.,2011; Cecere et al., 2013). Intriguingly, ZFP-1 and DOT-1.1 ChIP-chip peaks significantly overlap with O-GlcNAc ChIP-chip peaks at TSS of active genes. Although our extensive analyses determined that ZFP-1 and DOT1.1 do not carry this modification, we have found that the level of O-GlcNAc on chromatin is elevated in zfp-1 mutant worms, which correlates with the increased transcription of ZFP-1 target genes. On the other hand, we have determined that promoter-localized O-GlcNAc stimulates transcription of the same genes. These results indicate that ZFP-1 and O-GlcNAc have opposite roles in regulating transcription of metabolic genes. We are currently focusing on a candidate protein modified by O-GlcNAc and will present our progress in understanding its interplay with the ZFP-1/DOT-1.1 complex at the glucose-responsive promoters.

Ceballos, Ainhoa et al. (2013) International Worm Meeting "O-GlcNAcylation of a conserved chromatin factor, ZFP-1(AF10), as a possible glucose-sensing mechanism."

Ceballos, Ainhoa, Avgousti, Daphne, Alla, Grishok, Cecere, Germano

[International Worm Meeting, 2013]

The hallmark of Type 2 Diabetes is a defect in the glucose-sensing mechanism inducing insulin resistance. Here, we propose a novel epigenetic regulation of the insulin signaling pathway that involves a chromatin-associated protein and O-GlcNAcylation. Recently, we have described a role for C. elegans ZFP-1 protein, a conserved chromatin-binding factor, in negatively modulating transcription of the pdk-1 gene, which encodes a kinase critical for insulin signaling (Mansisidor et al., 2011). In addition, preliminary results from our laboratory show evidence of post-translational modifications of ZFP-1 and we have noticed almost identical genome localization profiles of ZFP-1 (Mansisidor et al., 2011) and O-GlcNAc modification (Love et al., 2010). O-GlcNAcylation is one of the most abundant eukaryotic post-translational modifications, which participates in the regulation of insulin production and modulates systems responsive to insulin. Based on these multiple observations, we are investigating whether ZFP-1 is modified by O-GlcNacylation, whether glucose stress promotes this modification and whether this leads to enhanced negative modulation of pdk-1 transcription. Finally, we are also interested in determining whether the mammalian homolog of ZFP-1, AF-10, is similarly involved in this new mechanism of insulin signaling inhibition.

Esse, R. et al. (2017) International Worm Meeting "The C. elegans DOT1L and c-Myc homologues cooperate in the nucleus to co-regulate target genes."

Ceballos, A., Grishok, A., Mora Martin, A., Gushchanskaia, E., Esse, R.

[International Worm Meeting, 2017]

Transcriptional control is mediated by interactions of transcription factors with their cognate DNA elements, as well as by epigenetic modifications to chromatin catalyzed by a variety of enzymes. Thus, understanding the crosstalk between transcription factors and epigenetic modifiers is of prime importance. The Dot1-like protein (DOT1L) is an evolutionary conserved methyltransferase with catalytic specificity towards histone 3 lysine 79 (H3K79). Owing to its crucial involvement in some aggressive leukemias, it has been gaining prominence in cancer research, with chemical inhibitors of DOT1L currently in clinical trials. Recent observations suggest that the role of DOT1L in malignant transformation can be generalized to contexts beyond leukemia. For instance, DOT1L has been implicated in breast cancer progression, and this has been attributed to its cooperation with c-Myc. However, the mechanistic details underlying this association are unknown. Previous work in our lab has shown that DOT-1.1, the C. elegans DOT1L homologue, is recruited to chromatin by ZFP-1 (similarly to DOT1L recruitment by AF10 in mammals), and this complex negatively modulates transcription. Interestingly, promoters of the ZFP-1/DOT-1.1 target genes are enriched in E-boxes, the consensus binding motif for c-Myc. Prompted by the exciting hypothesis that DOT-1.1 and MML-1, the C. elegans c-Myc homologue, cooperate genome-wide, we performed ChIP-seq for MML-1 and defined target genes as those with peak(s) close to their transcription start sites. Interestingly, the majority of MML-1 targets are also ZFP-1/DOT-1.1 targets. In order to understand the relevance of such co-localization for gene expression, we profiled mRNA and miRNA in wild-type worms and dot-1.1, zfp-1, and mml-1 lof mutants by microarrays. We found significant overlaps between genes upregulated in the three mutants, and the same was observed for downregulated genes. A significant global increase of non-coding transcripts was observed in either mutant compared with wild-type. Therefore, ZFP-1/DOT-1.1 and MML-1 co-regulate both coding and non-coding genes and globally inhibit non-coding transcription, including that of pri-miRNA. Recently, C. elegans enhancers were annotated based on their characteristic histone modifications signature, ATAC-seq profile and functional experiments. Notably, ZFP-1 localizes to ~50% of intronic enhancers. We have selected two of the enhancers for ChIP-qPCR and MML-1 was enriched at both. Thus, ZFP-1/DOT-1.1 and MML-1 co-regulate both promoters and enhancers. This is consistent with the developmental phenotypes (e.g. HSN migration defects) shared by the zfp-1, dot-1.1 and mml-1 lof mutants. Further investigation is underway to uncover the mechanism of cooperation of ZFP-1/DOT-1.1 and MML-1.

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.

Tuck S (2011) Curr Biol "Extracellular vesicles: budding regulated by a phosphatidylethanolamine translocase."

Tuck S

[Curr Biol, 2011]

Recent work on a Caenorhabditis elegans transmembrane ATPase reveals a central role for the aminophospholipid phosphatidylethanolamine in the production of a class of extracellular vesicles.