Nature Biotechnology
22, 871 - 876 (2004)
Published online: 6 June 2004; | doi:10.1038/nbt979
Temperature-sensitive control of protein activity by conditionally splicing inteinsMartin P Zeidler1, 2, 3, Change Tan1, 3, Yohanns Bellaiche1, 2, Sara Cherry1, Sabine Häder2, Urte Gayko1, 2
& Norbert Perrimon11
Department of Genetics, Howard Hughes Medical Institute, Harvard Medical School, 77 Avenue Louis Pasteur, Boston, Massachusetts 02115, USA. 2
Present addresses: Department of Molecular Developmental Biology, Max Planck Institute for Biophysical Chemistry, Am Fassberg 11, D-37077 Göttingen, Germany (M.P.Z. & S.H.), Institut Curie, UMR 144, 12 rue Lhomond, 75005 Paris, France (Y.B.), Amgen Inc., One Amgen Center Drive, Thousand Oaks, California 91320, USA (U.G.). 3
These authors contributed equally to this work.
Correspondence should be addressed to Norbert Perrimon perrimon@receptor.med.harvard.eduConditional or temperature-sensitive (TS) alleles represent useful tools with which to investigate gene function. Indeed, much of our understanding of yeast has relied on temperature-sensitive mutations which, when available, also provide important insights into other model systems. However, the rarity of temperature-sensitive alleles and difficulty in identifying them has limited their use. Here we describe a system to generate temperature-sensitive alleles based on conditionally active inteins. We have identified temperature-sensitive splicing variants of the yeast Saccharomyces cerevisiae vacuolar ATPase subunit (VMA) intein inserted within Gal4 and transferred these into Gal80. We show that Gal80-inteinTS is able to efficiently provide temporal regulation of the Gal4/upstream activation sequence (UAS) system in a temperature-dependent manner in Drosophila melanogaster. Given the minimal host requirements necessary for temperature-sensitive intein splicing, this technique has the potential to allow the generation and use of conditionally active inteins in multiple host proteins and model systems, thereby widening the use of temperature-sensitive alleles for functional protein analysis.
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