The kinetics of pre-mRNA processing in living cells is poorly known, preventing a detailed analysis of the regulation of these reactions. Using tetracycline-regulated promoters we performed, during a transcriptional induction, a complete analysis of the maturation of two cellular mRNAs, those for LT-␣ and ␤-globin. In both cases, splicing was appropriately described by first-order reactions with corresponding half-lives ranging between 0.4 and 7.5 min, depending on the intron. Transport also behaved as a first-order reaction during the early phase of ␤-globin expression, with a nuclear dwelling time of 4 min. At a later time, analysis was prevented by the progressive accumulation within the nucleus of mature mRNA not directly involved in export. Our results further establish for these genes that (i) splicing components are never limiting, even when expression is induced in naive cells, (ii) there is no significant RNA degradation during splicing and transport, and (iii) precursor-to-product ratios at steady state can be used for the determination of splicing rates. Finally, the comparison between the kinetics of splicing during transcriptional induction and during transcriptional shutoff reveals a novel coupling between transcription and splicing.In eukaryotic cells pre-mRNA maturation is a complex process that can entail the removal of multiple introns before a mature transcript can be exported to the cytoplasm. Moreover, the fidelity of processing and the control of alternative pathways are crucial for the correct execution of a genetic program. While our knowledge of the biochemical processes behind splicing and transport has greatly increased over the last years, we still know very little about their kinetics in vivo. This lack is all the more serious since the current view of nuclear RNA processing is that of a large number of regulators competing for the same substrates. This stresses the importance of the local concentration of regulators but also of the kinetics of the corresponding reactions. In addition, it is usually assumed that the RNA species generated during pre-mRNA processing are very unstable (7,21). If this is true, the level of expression should be critically dependent on the kinetics of processing.Our lack of knowledge of the kinetics of splicing and transport reflects the paucity of adequate experimental approaches. In vivo labeling studies have been used to investigate mRNA metabolism in mammalian cells. While this approach should provide access to the processing rates of pre-mRNA, two factors seriously limit its usefulness. (i) Studies on the fate of a specific RNA are hampered by the lack of sensitivity. (ii) The equilibration of the labeled compound with the intracellular nucleotide pool requires several minutes, which is the time scale anticipated for splicing and transport reactions. Thus the labeling is too slow for a stop flow type of kinetic analysis and too rapid for an approach to equilibrium analysis. Consequently, only some general information on pre-mRNA processing can be extr...

In the Drosophila bristle lineage, five differentiated cells arise from a precursor cell after a rapid sequence of asymmetric cell divisions (one every 2 hours). We show that, in mitotic cells, this rapid cadence of cell divisions is associated with cell cycles essentially devoid of the G1-phase. This feature is due to the expression of Cyclin E that precedes each cell division, and the differential expression of the S-transition negative regulator, Dacapo. Thus, apart from endocycles (G/S), which occurred in two out of five terminal cells, two other cell cycles coexist in this lineage: (1) an atypical cell cycle (S/G2/M), in which the S-phase is initiated during the preceding telophase; and (2) a canonical cell cycle (G1/S/G2/M) with a brief G1 phase. These two types of cell cycle result from either the absence or very transient expression of Dap, respectively. Finally, we show that the fate determinant factor, Tramtrack, downregulates Cyclin E expression and is probably involved in the exit of the cells from the cell cycle.

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