Joseph A. Curran and Benjamin Weiss (2016), What Is the Impact of mRNA 5' TL Heterogeneity on Translational Start Site Selection and the Mammalian Cellular Phenotype?, in Frontiers in Genetics (RNA)
, 7, 156.
Weiss B and Curran J. (2015), CAP+ selection: A combined chemical-enzymatic strategy for efficient eukaryotic mRNA enrichment via the 5’ cap., in Analytical Biochemistry
, 484, 72-74.
Legrand N. Araud T. Conne B. Kuijpers O. Jaquier-Gubler P and Curran J. (2014), An AUG codon conserved for protein function rather than translational initiation: the story of the protein sElk1., in PLoS ONE
, 9, e102890.
Rahim G. Araud T. Jaquier-Gubler P and Curran J. (2012), Alternative Splicing within the elk-1 5' Untranslated Region Serves To Modulate Initiation Events Downstream of the Highly Conserved Upstream Open Reading Frame 2., in Molecular and Cellular Biology
, 32, 1745-1756.
Dieudonné Francois-Xavier, O’Connor Patrick B.F, Gubler-Jaquier Pascale, Yasrebi Haleh, Conne Beatrice, Nikolaev Sergey, Antonarakis Stylianos, Baranov Pavel V, Curran Joseph, The Effect of Heterogeneous Transcription Start Sites (TSS) on the Translatome: Implications for the Mammalian Cellular Phenotype., in BMC Genomics
Legrand N. Jaquier-Gubler P and Curran J., The impact of the phosphomimetic eIF2αS/D on global translation, reinitiation and the integrated stress response is attenuated in N2a cells., in Nucleic Acids Research
The proposed project focuses on the regulation of protein synthesis and how this impacts on cellular physiology. It is very much a continuation and extension of the labs previous FNS grant. Recently, we have begun to focus on 5’UTR heterogeneity and its role in modulating the protein readout. Such variation arises as a result of alternative splicing and/or alternative promoter usage and serves to couple nuclear events to protein expression. These changes can have both quantitative (in terms of the amount of primary protein product synthesised) and qualitative (with regards to the types of primary protein products expressed) consequences. This heterogeneity may also “fine-tune” the expression of a particular gene to rapid intracellular changes. Despite the fact that as much as 22% of mammalian genes have alternative 5’UTRs, many of which are highly conserved, little is known about how this impacts on the cellular proteome. Furthermore, it remains unclear as to what extent non-conserved elements in the 5’UTR serve to modulate the translational readout in a species specific manner. Our work addresses these problems by focusing on a particular cellular gene that exhibits 5’ heterogeneity, namely, elk1. ELK-1 belongs to the subfamily of TCF transcription factors (Ternary Complex Factor) whose activation triggers the immediate early transcriptional response and the activation of genes such as c-fos. In rat neuronal tissue, a shorter N-terminally truncated protein, referred to as sELK-1, was reported. It is initiated from the 7th AUG start site on the mRNA and its expression correlated with NGF-driven neuronal differentiation and neurite outgrowth. Our lab has previously characterised two alternatively spliced 5’ UTR isoforms (5’UTRL and 5’UTRS) of the human gene. Both contained elements that play a role in translational regulation including two uAUGs (the second starting an ORF of only 2 codons) and stable RNA structures. The alternatively spliced exon is positioned just upstream of uORF2. Our published work indicated that leaky scanning, shunting and uORF2-mediated reinitiation all served to regulate ribosomal access to the AUGELK-1 (the 3rd on the mRNA). It also demonstrated that in human cells all ribosomes detected downstream of the AUGELK-1 were in a delayed reinitiation mode. These ribosomes were sequestered by a series of highly conserved, out-of-frame, iAUGs that served as repressors for sELK-1 expression in this cellular context. Recently, we have recently characterised the 5’UTRs from rat (PC12 cell line) and mouse (the N2a neuroblastoma cell line) and observed that the alternatively spliced exonII is species-specific. Furthermore, despite conservation of the iAUGs in these species we readily observed sELK-1 protein expression in N2a and differentiated PC12 cells. Our results, based upon the human system, suggests that this may arise due to a de-repression of negative structural elements within the 5’UTRs (possibly associated with the expression of RNA helicases) coupled to changes in ternary complex levels. They also point to differences in the behaviour of the alternatively spliced transcripts. In this proposal we aim to test this model.