Cultures of Escherichia coli (strains ML30 and K12 AB1157), synchronized by repeated phosphate starvation, were submitted to nutritional shifts-up at various cell ages. The progression of the replication forks was assessed by DNA-DNA hybridization of pulse-labelled chromosomal DNA with plasmid DNA probes containing specific chromosomal sequences. The rate of phospholipid synthesis and its cyclic discontinuities were measured by continuous and pulse labelling with palmitate. The DNA-DNA hybridization experiments showed that a shiftup induces a burst of initiation from the oriC region. These hybridization results, taken together with older data from the literature, suggest that most DNA initiations belonging to this burst are not followed by complete replication. Following a shift-up, the rate of phospholipid synthesis is maintained for 13 -20 min, depending on cell age at shift-up, then doubles. The new steady-state rate of phospholipid synthesis is reached through a series of three doublings, while the cell mass doubles approximately twice. This discrepancy brings the rate of phospholipid synthesis per mass unit to its steady-state postshift value.Bacteria are able to grow in a constant environment for numerous generations without any variation in their chemical composition or internal organization. The biochemical analysis of the cyclic events taking place in such cells has become possible through the synchronization of large populations. However, the study of synchronous cultures growing in a constant environment does not allow one to deduce the interrelationships between the discontinuous cyclic events of bacterial life, save a description of the temporal relationships between these events. On the other hand, nutritional shifts have long been considered as tools to approach cell cycle regulation [l]. When Escherichia coli cells are shifted from a nutritionally poor to a rich medium, ribosome synthesis reaches its postshift value almost immediately [2, 31 and the bacterial growth rate quickly increases [l, 41. Among the known cycle-related events, cell division and DNA replication following a shift-up have been much studied. The classical view is that, after a shift-up, these two parameters show a 'rate maintenance' during about 60 min for cell division [5 -91 and about 20 min for DNA replication [5 -7, 101 at which time their rates increase abruptly to their postshift values. As far as the effects of a shift-up on cell division are concerned, a first experimental improvement was brought about by using synchronous populations [l 1, 121 and a second improvement arose from the use of nutritional pulses (shifts-up followed after less than 15 min by shifts-down back to the original poor medium) in synchronous populations [12]. We showed byCorrespondence to F. KCpts, c/o R. Schekman, Department of Biochemistry, University of California, Berkeley, California USA-94720This article is dedicated to the memory of the late Dr Adam Kepis.Abbreviation. DROPS, doubling of the rate of phospholipid synthesis.~~ these means th...
