BackgroundPredictable control of gene expression is necessary for the rational design and optimization of cell factories. In the yeast Saccharomyces cerevisiae, the promoter is one of the most important tools available for controlling gene expression. However, the complex expression patterns of yeast promoters have not been fully characterised and compared on different carbon sources (glucose, sucrose, galactose and ethanol) and across the diauxic shift in glucose batch cultivation. These conditions are of importance to yeast cell factory design because they are commonly used and encountered in industrial processes. Here, the activities of a series of “constitutive” and inducible promoters were characterised in single cells throughout the fermentation using green fluorescent protein (GFP) as a reporter.ResultsThe “constitutive” promoters, including glycolytic promoters, transcription elongation factor promoters and ribosomal promoters, differed in their response patterns to different carbon sources; however, in glucose batch cultivation, expression driven by these promoters decreased sharply as glucose was depleted and cells moved towards the diauxic shift. Promoters induced at low-glucose levels (PHXT7, PSSA1 and PADH2) varied in induction strength on non-glucose carbon sources (sucrose, galactose and ethanol); in contrast to the “constitutive” promoters, GFP expression increased as glucose decreased and cells moved towards the diauxic shift. While lower than several “constitutive” promoters during the exponential phase, expression from the SSA1 promoter was higher in the post-diauxic phase than the commonly-used TEF1 promoter. The galactose-inducible GAL1 promoter provided the highest GFP expression on galactose, and the copper-inducible CUP1 promoter provided the highest induced GFP expression following the diauxic shift.ConclusionsThe data provides a foundation for predictable and optimised control of gene expression levels on different carbon sources and throughout batch fermentation, including during and after the diauxic shift. This information can be applied for designing expression approaches to improve yields, rates and titres in yeast cell factories.Electronic supplementary materialThe online version of this article (doi:10.1186/s12934-015-0278-5) contains supplementary material, which is available to authorized users.
Chinese hamster ovary (CHO) cells are the predominant host cell line for the production of biopharmaceuticals, a growing industry currently worth more than $188 billion USD in global sales. CHO cells undergo programmed cell death (apoptosis) following different stresses encountered in cell culture, such as substrate limitation, accumulation of toxic by‐products, and mechanical shear, hindering production. Genetic engineering strategies to reduce apoptosis in CHO cells have been investigated with mixed results. In this review, a contemporary understanding of the real complexity of apoptotic mechanisms and signaling pathways is described; followed by an overview of antiapoptotic cell line engineering strategies tested so far in CHO cells.
The authors conclude that CSD elicited by CCI can mask the true effect of TBI on hypercapnic vasodilation for at least 30 minutes. Exogenous NO, but not papaverine, can reverse the attenuation of cerebrovascular reactivity to CO2 caused by TBI. This result supports the hypothesis that NO production is reduced after TBI and that the NO donor has a potential beneficial role in the clinical management of head injury.
Various combinations of cranial remodeling techniques are used in an attempt to provide optimal cosmetic results and to reduce possible sequelae associated with craniosynostosis. One element of deformity that is difficult to correct directly is an overly flattened area such as that found in the parietal area in sagittal synostosis, unilaterally in lambdoid synostosis, or even in severe positional molding. The authors present a novel application for recontouring cranial bone, namely the multiple-revolution spiral osteotomy. The advantages of this technique include the avoidance of large areas of craniectomy and immediate correction of the cranial deformity. The surgical procedure, illustrative cases, early results, and apparent benefits of this technique are discussed.
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