Loss of cell viability, through engagement of apoptotic cell death, represents a limitation to maintenance of high levels of productivity of recombinant animal cells in culture. The ability to monitor the status of recombinant cells, and to define indicators of their "well-being," would present a valuable approach to permit a rational intervention at appropriate times during culture. Growth arrest and DNA damage gene 153 (GADD153) is a member of the CCAAT/enhancer-binding protein (C/EBP) family of transcription factors and has been associated with apoptosis. We have examined the expression of GADD153 in conditions associated with apoptosis of recombinant CHO cells in batch culture. GADD153 expression is very low in CHO cells growing in the exponential phase of batch culture but is activated as cells enter the decline phase. Depletion of nutrients (glucose or glutamine) causes activation of GADD153 expression as does the imposition of endoplasmic reticulum stress. In all cases, there is a good relationship between the extent of apoptosis that occurs in response to each stress and the degree of GADD153 expression. In addition, nutrient refeeding or reversal of stress produces a concomitant decrease in expression of GADD153 and the susceptibility to apoptosis. Thus, GADD153 appears to offer a valid indicator of apoptosis and illustrates the potential for definition of monitors of cellular status related to the likelihood of apoptosis of cell populations.
A bacterium that utilized propane as a sole carbon source was isolated from soil and identified as a strain of Rhodococcus rhodochrous. Of the n-alkanes tested ((2,-C,) it grew only on propane, and it was not capable of growth on alkenes. The organism grew on most of the potential intermediates of propane metabolism and simultaneous adaptation studies showed that it could oxidize both terminal and sub-terminal intermediates. Assays of enzyme activities in cell-free extracts revealed elevated levels of enzymes of both terminal and sub-terminal pathways in propane-grown cells. The initial propane-specific oxygenase activity was measured by its ability to co-oxidize propene to epoxypropane. This oxygenase system was investigated in terms of its inhibitor profile and was compared with n-alkane oxygenase systems described in the literature. On the basis of the comparison, the oxygenase appears to be of a type not previously reported.
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