Diviak, Bridget et al. (2021) International Worm Meeting "Characterization of the Role of the Terminal Adenosine Located at the pre-mRNA Cleavage Site"

Kendall, Carleigh, O'Brien, Shannon, Mangone, Marco, Heyza, Mairead, Diviak, Bridget

[International Worm Meeting, 2021]

The region downstream of the STOP codon in mRNA, referred to as the 3'Untranslated Region (3'UTR), governs the length of mature mRNA. Specifically, the cleavage site located in this region determines where mRNA cleavage will occur and where polyadenylation reaction will begin, thus terminating mRNA transcription. The mRNA cleavage and polyadenylation machinery in C. elegans is highly conserved to its human counterpart, with most functional domains and critical amino acids preserved. Dysregulation of 3'UTR processing has been observed in many diseases, such as cancer, Alzheimer's disease, and muscular dystrophies, but unfortunately the molecular mechanisms underlying the mRNA transcription termination remain elusive. Although the exact cleavage site is not precise, our lab has identified an adenosine consistently located at the mRNA cleavage site. It is unclear if this adenosine is maintained in the mature mRNA transcripts proceeding cleavage and/or is used as a template for the polymerization of the poly(A) tail. In order to answer this question, we developed a novel terminal adenosine RNA methyltransferase (TAM) assay that will sense the inclusion or exclusion of this terminal adenosine at the cleavage site of C. eleganstranscripts by taking advantage of the human nuclear methyltransferase, METTL16. METTL16 methylates the underscored adenosine in its binding motif, "UACAGAGAA", in both mRNA and snRNA. We have cloned both the human METTL16 gene and its RNA recognition motif at the cleavage site of the C. elegans gene M03A1.3and co-expressed them both in the pharynx tissue. Understanding this process is crucial to identifying the main mechanisms behind mRNA cleavage site determination, further advancing knowledge in gene regulation which influences development, growth, and disease.

David R. Bell et al. (2006) European Worm Meeting "The Function of Latrophilin in C. elegans"

S. van der Poel, P.N.R. Usherwood, D. Kendall, J. Davy, A. Ademola, I.R. Duce, D. de Pomerai, David R. Bell, I. R. Mellor

[European Worm Meeting, 2006]

David R. Bell, A. Ademola, J. Davy, S. van der Poel, D. Kendall, P.N.R. Usherwood, I.R. Duce, I. R. Mellor and D. de Pomerai. Black Widow spider venom (BWSV) contains latrotoxins that induce catastrophic neurotransmitter release in mammals, and is known to bind with high affinity to three neural proteins in mammals, including latrophilin. We have investigated C. elegans as a model system for studying the function of these proteins, and their role in regulating neurotransmitter release by latrotoxins.. We have shown that latrophilin is required for the lethality of BWSV in C. elegans by RNAi experiments. We now present data on the characterisation of null mutants of the C. elegans latrophilin, lat-1, illustrating the function of this gene in C. elegans, and show that lat-1 nulls are resistant to emodepside. The expression of latrophilin-GFP constructs is examined, and compared with the phenotypes seen in null mutants. Genetic interactions between endogenous signaling pathways and the lat-1 nulls have been examined by the construction of double mutant worms, revealing novel facets of lat-1 function.

Catherine M Dempsey et al. (2005) International Worm Meeting "A genetic screen for the targets of Fluoxetine in the egg-laying paradigm."

Catherine M Dempsey, Ji Y Sze

[International Worm Meeting, 2005]

There is a crisis of confidence in one of the most widely used psychiatric drugs, selective serotonin (5-HT) re-uptake inhibitors (SSRIs). Thought to boost synaptic levels of 5-HT by blocking its re-absorption from the synaptic cleft via the 5-HT re-uptake transporter (SERT), SSRIs were widely touted as the most effective treatment for depression in mature patients. Recent reports show that SSRI exposure during key developmental stages may increase the risk of depression and suicide in both mice and humans (1,2). In fact, even in adults some studies suggest a twofold increase in the rate of suicide attempts in patients receiving SSRIs compared with placebo or therapeutic interventions other than tricyclic antidepressants (3). Despite such misgivings the genetic basis and molecular mechanisms of the most widely used antidepressant in history Prozac (fluoxetine) remain largely unknown. Fluoxetine and 5-HT both stimulate egg-laying in C. elegans. Fluoxetines action is both 5-HT dependent and independent, it can still induce, at least partially, egg-laying in both 5-HT (tph-1 (mg280)) and SERT (mod-5 (n3314)) deficient animals. To distinguish the action of 5-HT from that of fluoxetine we are conducting a genetic screen for resistance to fluoxetine stimulation of egg-laying in a ser-1 (ok345) mutant background. SER-1 is a G-protein coupled receptor expressed in the vulval muscles; mutants lacking this gene are fully resistant to 5-HT stimulated egg-laying but respond substantially to fluoxetine (4). F2 progeny of ethylmethane sulfonate mutagenized animals are exposed to 0.5 mg/ml fluoxetine and resistant animals who lay no eggs are recovered. Egg-laying can also be stimulated by the tricyclic antidepressant imipramine via a different genetic pathway which includes the 5-HT receptor SER-4 (4). To select specifically fluoxetine resistant mutants, we compare egg-laying rates of mutants in fluoxetine and imipramine, animals capable of laying eggs in the presence of imipramine but not fluoxetine are studied further. 1. Whittington, C. J., Kendall, T., Fonagy, P., Cottrell, D., Cotgrove, A. & Boddington, E.(2004) Lancet 363, 1341-1345. 2. Ansorge, M. S., Zhou, M. M., Lira, A., Hen, R. & Gingrich, J. A. (2004) Science 306, 879-881.3. Fergusson, D., Doucette, S., Cranley, K., Glass, K. C., Shapiro, S., Healy, D., Hebert, P. & Hutton, B. (2005) British Medical Journal 330, 396-399.4. Dempsey, C. M. Mackenzie S. M., Gargus A., Blanco G. & Sze J. Y. (2005). 5-HT (5HT), Genetics (In press)