Development of a novel simplified Q-system with integrated temporal expression control

Abstract

Binary gene expression systems have played a major role in uncovering the function of genes, cells and organ systems during and after development. Employed initially in model systems such as flies and mice, advances in gene technology have vastly expanded the number of species in which these systems can be deployed. One of their limitations is the challenge of imposing temporal expression control. Here, we report the incorporation of temperature-sensitive intein modules with different temperature profiles into QF2, the transcription factor anchoring the Q-system widely used in Drosophila and introduced recently into several insect species and the zebrafish D. rerio. Intein removal from temperature-sensitive QF2_INTts activators in Drosophila larvae and adult flies raised under permissive conditions (18 degree Celsius to 25 degree Celsius) renders QF2 active and drives strong expression of QUAS reporters. In contrast, raising Drosophila at restrictive temperatures (25 degree Celsius to 30 degree Celsius) keeps QF2_INTts in a non-functional state unable to bind DNA and therefore keeping QUAS reporters inactive. We further show that reporter expression can be turned on and off both in larvae and flies by changing temperature conditions between restrictive to permissive. Finally, expression of the inward rectifying potassium channel (QUAS-Kir2.1) under the control of QF2_INTts drivers in olfactory neurons and pan-neuronally was effectively suppressed in flies raised under restrictive temperature conditions with no impact on olfactory acuity or survival, respectively. In contrast, when respective flies were shifted to permissive temperature-condition, flies lost olfactory acuity and became paralyzed within one to four days, respectively. Thus, the intein-based Q-system, henceforth referred to as QINT system, renders the difficult to employ drug-dependent suppressor QS unnecessary, greatly facilitating temporal regulation of gene expression. Having several advantages over the GAL4 system, the Q system has gained broad acceptance in other insect species and the zebrafish D. rerio, and thus the temperature-regulatable QINT system can be implemented in many other organisms, including disease-transmitting mosquitoes and poikilotherm vertebrate animals to study gene and cell function at any time during or after development.