Analysis of neuropeptide gene expression by transfection of DNA into cell lines

The transcriptional regulation of neuropeptide genes by cAMP is often directed by a cAMP responsive enhancer (CRE) upstream to the promoter of the genes. The identity of the CRE was determined by transient transfection experiments and has the consensus sequence of 5′-TGACGTCA-3′. A large family of transcription factors have been identified which recognize the CRE. Transient transfection assays that employed expression of these factors driven by viral promoters have determined they can transactivate the neuropeptide gene promoters. Because of the large number of factors that have been identified as being able to recognize the CRE, it has been difficult to determine which factors mediate in vivo the transactivation of a particular CRE. Using a dominant negative mutant of one of these factors, CREB, it has been determined that both CREB as well as other factors which do not interact with CREB may mediate the cAMP response of the somatostatin gene.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/43232/1/11022_2005_Article_BF01667368.pd

Reduction of translating ribosomes enables Escherichia coli to maintain elongation rates during slow growth

Bacteria growing under different conditions experience a broad range of demand on the rate of protein synthesis, which profoundly affects cellular resource allocation. During fast growth, protein synthesis has long been known to be modulated by adjusting the ribosome content, with the vast majority of ribosomes engaged at a near-maximal rate of elongation. Here, we systematically characterize protein synthesis by Escherichia coli, focusing on slow-growth conditions. We establish that the translational elongation rate decreases as growth slows, exhibiting a Michaelis-Menten dependence on the abundance of the cellular translational apparatus. However, an appreciable elongation rate is maintained even towards zero growth, including the stationary phase. This maintenance, critical for timely protein synthesis in harsh environments, is accompanied by a drastic reduction in the fraction of active ribosomes. Interestingly, well-known antibiotics such as chloramphenicol also cause a substantial reduction in the pool of active ribosomes, instead of slowing down translational elongation as commonly thought.NIH [R01GM109069, R01GM072528, GM118850]; National Natural Science Fund of the People's Republic of China (NSFC) [31530081]; China Scholarship Council (CSC) [201306010039]SCI(E)ARTICLE2