Rapid generation of hypomorphic mutations

Arthur, Laura L.; Chung, Joyce J.; Jankirama, Preetam; Keefer, Kathryn M.; Kolotilin, Igor; Pavlovic-Djuranovic, Slavica; Chalker, Douglas L.; Grbić, Vojislava; Green, Rachel; Menassa, Rima; True, Heather L.; Skeath, James B.; Djuranović, Sergej

doi:10.1038/ncomms14112

Cited by 17 publications

(27 citation statements)

References 66 publications

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“…As expected, we observed robust changes in mRNA levels ( Figure 3A) and substantial losses in protein expression ( Fig. 3B) for reporters with polyA tracks in both neonatal human dermal fibroblasts (HDFs) and AT-genome rich T. thermophila (10,11). We observed minimal, if any effects, of polyA track insertion on reporter mRNA and protein expression in P. falciparum (Fig.…”

Section: Resultssupporting

confidence: 81%

“…Recently, it was demonstrated that the translation of genes with polyadenosine runs (polyA tracks), primarily coding for lysine residues, is attenuated in the majority of tested eukaryotic and prokaryotic organisms due to ribosomal stalling and frameshifting on such RNA motifs (8)(9)(10)(11)(12)(13)(14)(15). In humans, the presence of just 12 adenosines in an mRNA coding region will deplete the protein synthesis of a given gene by more than 40% (9,10). The consequence of translational arrest is activation of one or more mRNA surveillance mechanisms and has been demonstrated in studies of human, yeast and E. coli (9,12).…”

Section: Mainmentioning

confidence: 99%

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PolyA tracks and poly-lysine repeats are the Achilles heel ofPlasmodium falciparum

Sp¹,

Erath²,

Andrews

et al. 2018

Preprint

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Plasmodium falciparum, the causative agent of human malaria, is an apicomplexan parasite with a complex, multi-host life cycle. Sixty percent of transcripts from its extreme AT-rich (81%) genome possess coding polyadenosine (polyA) runs, distinguishing the parasite from its hosts and other sequenced organisms. Recent studies indicate that transcripts with polyA runs encoding poly-lysine are hot spots for ribosome stalling and frameshifting, eliciting mRNA surveillance pathways and attenuating protein synthesis in the majority of prokaryotic and eukaryotic organisms. Here, we show that the P. falciparum translational machinery is paradigm-breaking. Using bioinformatic and biochemical approaches, we demonstrate that both endogenous genes and reporter sequences containing long polyA runs are efficiently and accurately transcribed and translated in P. falciparum cells. Translation of polyA tracks in the parasite does not elicit any response from mRNA surveillance pathways usually seen in host human cells or organisms with similar AT content. The translation efficiency and accuracy of the parasite protein synthesis machinery reveals a unique role of ribosomes in the evolution and adaptation of P. falciparum to an AU-rich transcriptome and polybasic amino sequences. Finally, we show that the ability of P. falciparum to synthesize long poly-lysine repeats has given this parasite a unique protein exportome and an advantage in infectivity that can be suppressed by addition of exogenous poly-basic polymers.

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Section: Resultssupporting

confidence: 81%

Section: Mainmentioning

confidence: 99%

PolyA tracks and poly-lysine repeats are the Achilles heel ofPlasmodium falciparum

Sp¹,

Erath²,

Andrews

et al. 2018

Preprint

Self Cite

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show abstract

“…In this context, rather than slowing elongation, the reduced mRNA structure caused by AT-rich nonoptimal codons is suggested to enhance the rate of translation and increase translation efficiency, possibly by optimizing the spacing between ribosomes (40). Yet, these nonoptimal codons might also impact the efficiency of translation initiation, as including more than eight consecutive nonoptimal codons in the 5Ј end of coding sequences causes decreased protein expression (91,92).…”

Section: Ribosome Stalling Dictates Gene Expressionmentioning

confidence: 99%

The stop-and-go traffic regulating protein biogenesis: How translation kinetics controls proteostasis

Stein

Frydman

2019

Journal of Biological Chemistry

128

122

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“…Subsequently, to test whether RhoGAPp190 mRNA upregulation can contribute to the locomotor dysfunction observed in the tbph-null flies, we assayed the genetic interaction between RhoGAPp190 and tbph by examining the consequences of the combined RhoGAPp190 / tbph silencing on climbing ability. Since flies bearing homozygous tbph-null mutations show limited larval lethality [ 15 – 18 ], tbph hypomorphic flies (carrying a milder phenotype) [ 19 – 22 ] were used to test the genetic interaction between this nuclear factor and RhoGAPp190. Two different RhoGAPp190 RNAi fly lines (namely, 6429 and 6430), generated by expressing RhoGAPp190 and tbph RNAi in tbph hypomorphic flies, were used for this experiment.…”

Section: Resultsmentioning

confidence: 99%

RhoGAPp190: A potential player in tbph-mediated neurodegeneration in Drosophila

et al. 2018

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TDP-43 is an ubiquitous and highly conserved ribonucleoprotein involved in several cellular processes including pre-mRNA splicing, transcription, mRNA stability and transport. Notwithstanding the evidence of TDP-43 involvement in the pathogenesis of different neurodegenerative disorders (i.e. ALS and FTLD), the underlying mechanisms are still unclear. Given the high degree of functional similarity between the human and fly orthologs of TDP-43, Drosophila melanogaster is a simple and useful model to study the pathophysiological role of this protein in vivo. It has been demonstrated that the depletion of the TDP-43 fly ortholog (tbph) induces deficient locomotive behaviors and reduces life span and anatomical defects at the neuromuscular junction. In this study, using the known binding specificity of TDP-43/tbph for (UG) repeated sequences, we performed a bioinformatic screening for fly genes with at least 6 (TG) repeats in a row within the 3'-UTR regions in order to identify the genes that might be regulated by this factor. Among these genes, we were able to identify RhoGAPp190 as a potential target of the tbph-mediated neurodegeneration. RhoGAPp190 is a negative regulator of Drosophila RhoA, a GTPase protein implicated in the fine modulation of critical cellular processes including axon branch stability and motor axon defasciculation at muscle level and cognitive processes. We were able to demonstrate that the RhoGAPp190 expression is upregulated in a tbph-null fly model, providing evidence that this deregulation is associated to tbph silencing. Our results introduce RhoGAPp190 as a novel potential mediator in the complex scenario of events resulting from in vivo tbph loss-of-function.

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Rapid generation of hypomorphic mutations

Cited by 17 publications

References 66 publications

PolyA tracks and poly-lysine repeats are the Achilles heel ofPlasmodium falciparum

PolyA tracks and poly-lysine repeats are the Achilles heel ofPlasmodium falciparum

The stop-and-go traffic regulating protein biogenesis: How translation kinetics controls proteostasis

RhoGAPp190: A potential player in tbph-mediated neurodegeneration in Drosophila

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