Mendonsa S., von Kuegelgen N., Bujanic L. and Chekulaeva M. (2021). Charcot-Marie-Tooth mutation in glycyl-tRNA synthetase stalls ribosomes in a pre-accommodation state and activates integrated stress response. Nucleic Acids Research, https://doi.org/10.1093/nar/gkab730
Toxic gain-of-function mutations in aminoacyl-tRNA synthetases cause neurodegeneration, known as Charcot–Marie–Tooth (CMT) disease. Here we use high-resolution ribosome profiling and interaction studies to show that CMT mutations in glycyl-tRNA synthetase deplete the pool of glycyl-tRNAᴳˡʸ available for translation and cause ribosomes to pause at glycine codons, which in turn activates the integrated stress response (ISR). Thus, providing a supply tRNAGly and inhibition of ISR emerge as therapeutic strategies to alleviate degeneration in CMT.
Ludwik K.A., von Kuegelgen N. and Chekulaeva M. (2019). Genome-wide analysis of RNA and protein localization and local translation in mESC-derived neurons. Methods issue on experimental and computational techniques for studying structural dynamics and function of RNA, http://dx.doi.org/10.1016/j.ymeth.2019.02.002
von Kuegelgen N. and Chekulaeva M. (2020). Conservation of a core neurite transcriptome across neuronal types and species. WIREs RNA https://www.ncbi.nlm.nih.gov/pubmed/32059075
Recent high-throughput analyses have revealed that neurites contain hundreds to thousands of mRNAs, but an analysis comparing the transcriptomes derived from these studies has been lacking. Here we analyze 20 datasets pertaining to neuronal mRNA localization across species and neuronal types and identify a conserved set of mRNAs that had robustly localized to neurites in a high number of the studies. The set includes mRNAs encoding for ribosomal proteins and other components of the translation machinery, mitochondrial proteins, cytoskeletal components, and proteins associated with neurite formation. Our combinatorial analysis provides a unique resource for future hypothesis-driven research. Check lab resources for the interactive table that allows you to search to localization, expression and translation patterns of your favourite transcripts across 20 neuronal datasets.
Ciolli Mattioli C., Rom A., Franke V., Imami K., Arrey G., Terne M., Woehler A., Akalin A., Ulitsky I., and Chekulaeva M. (2018). Alternative 3′ UTRs direct localization of functionally diverse protein isoforms in neuronal compartments. Nucleic Acids Research, https://doi.org/10.1093/nar/gky1270
RNA localization is mediated by specific cis-regulatory elements usually found in mRNA 3'UTRs. Therefore, processes that generate alternative 3'UTRs – alternative splicing and polyadenylation – have the potential to diversify mRNA localization patterns in neurons. Here we showed that usage of alternative 3'UTRs serves as a novel mechanism mediating localization of functionally distinct protein isoforms to different subcellular compartments.
Chekulaeva M. and Rajewsky N. (2018). Roles of Long Noncoding RNAs and Circular RNAs in Translation. Cold Spring Harbor Perspectives in Biology, https://doi.org/10.1101/cshperspect.a032680
Zappulo, A.*, van den Bruck, D.*, Ciolli Mattioli, C.*, Franke, V.*, Imami, K., McShane, E., Moreno-Estelles, M., Calviello, L., Filiipchyk, A., Peguero-Sanchez, E., Mueller, T., Woehler, A., Birchmeier, C., Merino, E., Rajewsky, N., Ohler. U., Mazzoni, E., Selbach, M., Akalin, A., and Chekulaeva, M. (2017). RNA localization is a key determinant of neurite-enriched proteome. Nature Communications, doi: http://dx.doi.org/10.1038/s41467-017-00690-6
The subcellular localization of proteins is fundamental to neuronal growth and synaptic plasticity which is the basis of learning and memory. Protein localization can be achieved (1) by transporting proteins as parts of RNPs or vesicular organelles; (2) through mRNA localization and local translation; or (3) via local translation of equally distributed mRNAs. While specific examples for these mechanisms have been described in the literature, it was unclear how much each of these mechanisms contributes to the overall asymmetry of protein distribution in neurons. To fill in this gap, we developed neuron fractionation scheme in combination with mass spectrometry, RNAseq, Riboseq and bioinformatics analyses to identify proteins and RNAs that are differentially localized and translated between neurites and soma. This analysis resulted in two key messages: (1) almost half on the neurite-enriched proteome is encoded by neurite-localized mRNAs, revealing mRNA localization as a key determinant of protein localization to neurites; (2) we generated a unique resource of local neuronal transcriptome, proteome and translatome and identified dozens of neurite-targeted non-coding RNAs and RNA-binding proteins with potential regulatory roles in neuronal polarity.
Pamudurti, N.R.*, Bartok, O.*, Jens, M.*, Ashwal-Fluss, R.*, Stottmeister, C., Ruhe, L., Hanan. M., Wyler, M., Perez-Hernandez, D., Ramberger, E., Shenzis, S., Samson, M., Dittmar, G., Landthaler, M., Chekulaeva, M., Rajewsky, N. and Kadener, S. (2017). Translation of circRNAs. Molecular Cell, doi: http://dx.doi.org/10.1016/j.molcel.2017.02.021
Mauri, M., Kirchner, M., Aharoni, R., Mattioli, C., van den Bruck, D., Gutkovitch, N., Modepalli, V., Selbach, M., Moran, Y., and Chekulaeva, M. (2016). Conservation of miRNA-mediated silencing mechanisms across 600 million years of animal evolution. Nucleic Acids Research, doi: 10.1093/nar/gkw792
Our current knowledge about the mechanisms of miRNA silencing is restricted to few lineages such as vertebrates, arthropods, nematodes and land plants. In our earlier work we have shown that miRNA-mediated silencing in bilaterian animals is dependent on the tryptophan-containing motifs (W-motifs) dispersed throughout the GW182 protein, the key components of the miRNP (Chekulaeva et al., 2011). Here, we dissect the function of GW182 protein in the cnidarian Nematostella, separated by 600 million years from other Metazoa. Using cultured human cells, we show that Nematostella GW182 recruits the CCR4-NOT deadenylation complexes via its W-motifs, thereby inhibiting translation and promoting mRNA decay. Thus, our work suggests that this mechanism of miRNA-mediated silencing was already active in the last common ancestor of Cnidaria and Bilateria.
Chekulaeva, M, Landthaler, M. (2016). Eyes on translation. Mol. Cell 63(6):918-925, doi: http://dx.doi.org/10.1016/j.molcel.2016.08.031
Translation is a key step of gene expression, regulated via multiple mechanisms. Here we review recent advances in quantification and visualization of translation, focusing on novel imaging approaches to study translation dynamics of single mRNAs in live cells.
Chekulaeva*, M., Mathys*, H., Zipprich, J., Attig, J., Colic, M., Parker, R., and Filipowicz, W. (2011). miRNA repression involves GW182-mediated recruitment of CCR4–NOT through conserved W-containing motifs. Nature Structural and Molecular Biology 18(11): 1218-26.
In this manuscript, we demonstrate that miRNA silencing is mediated by novel conserved linear motifs (W-motifs) dispersed throughout GW182, the effector protein of miRNA repression complex. These motifs recruit deadenylation complexes to cause deadeanylation, decay and translational repression of target mRNAs. This discovery reconciles literature data, which could not be explained by previous models and provides a new foundation from which to explore miRNA function.
Chekulaeva, M., Parker, R., and Filipowicz, W. (2010). The GW/WG repeats of GW182 function as effector motifs for miRNA-mediated repression. Nucleic Acids Research 6673-83.
Ozgur, S, Chekulaeva, M., Stoecklin, G. (2010). Human Pat1b connects deadenylation with mRNA decapping and controls the assembly of processing-bodies. Molecular and Cellular Biology 30(17): 4308-23.
Chekulaeva, M. and Filipowicz, W. (2009). Mechanisms of miRNA-mediated post-transcriptional regulation in animal cells. Current Opinion in Cell Biology 21(3): 452-60.
Chekulaeva, M., Filipowicz, W., and Parker, R. (2009). Multiple independent domains of dGW182 function in miRNA-mediated repression in Drosophila. RNA 15: 794-803.
Chekulaeva, M., Hentze, M.W., and Ephrussi, A. (2006). Bruno acts as a dual repressor of oskar mRNA translation promoting mRNA oligomerization and formation of silencing particles. Cell 124: 521-533.
In this work we uncovered a novel mechanism of translational repression that involves mRNA oligomerization into unusually large RNP complexes, silencing particles, that cannot be accessed by ribosomes. This mechanism is particularly suited to coupling translational control with mRNA transport, a common and ill-understood theme in developmental biology and neurobiology.
Chekulaeva, M., and Ephrussi, A. (2004). Drosophila Development: RNA Interference ab ovo. Current Biology 14: 428-30.
Chekulayeva M.N., Kurnasov O.V., Shirokov V.A., and Spirin A.S. (2001). Continuous-exchange cell-free protein-synthesizing system: synthesis of HIV-1 antigen Nef. Biochem Biophys Res Commun. 280(3):914-7.