Abstract
Secretion of an intracellular protein from a cell factory requires as a first step the redirection of the mRNA for synthesis of the protein on the endoplasmic reticulum. The feasibility of retargeting a mRNA coding for an intracellular protein to the endoplasmic reticulum was investigated using Ltk- fibroblasts stably transfected with gene constructs in which rabbit beta-globin coding region and 5'UTR was linked to albumin signal sequence and different 3'untranslated regions. Globin transcripts with the native globin 3'untranslated region or with the 3'untranslated region of c-myc are present in free/cytoskeletal-bound polysomes. The addition of the signal sequence from rat albumin redirects both these globin transcripts to membrane-bound polysomes but the presence of the c-myc 3'UTR reduces the extent of redirection. Globin transcripts with both the signal sequence and 3'untranslated region from the albumin gene are efficiently redirected to membrane-bound polysomes. The results suggest competition between 5' and 3' localising signals. The addition of the signal sequence does not destabilise the mRNA nor affect translational efficiency. It is concluded that it is possible to retarget an mRNA to the endoplasmic reticulum while maintaining stability and translational capacity. This has important implications for the development of vectors to promote secretion of intracellular proteins from cell factories.
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Partridge, K.A., Johannessen, A., Tauler, A. et al. Competition between the signal sequence and a 3'UTR localisation signal during redirection of beta-globin mRNA to the endoplasmic reticulum: implications for biotechnology. Cytotechnology 30, 37–47 (1999). https://doi.org/10.1023/A:1008079901508
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DOI: https://doi.org/10.1023/A:1008079901508