Cultivation of retrovirus packaging cells at 32˚C represents a common procedure to achieve high titres in mouse retrovirus production. Gene expression profiling of mouse NIH 3T3 cells producing amphotropic mouse leukaemia virus 4070A revealed that 10 % of the 1176 cellular genes investigated were regulated by temperature shift (37/32˚C), while 5 % were affected by retrovirus infection. Strikingly, retrovirus production at 32˚C activated the cholesterol biosynthesis/transport pathway and caused an increase in plasma membrane cholesterol levels. Furthermore, these conditions resulted in transcriptional activation of smoothened (smo), patched (ptc) and gli-1; Smo, Ptc and Gli-1, as well as cholesterol, are components of the Sonic hedgehog (Shh) signalling pathway, which directs pattern formation, diversification and tumourigenesis in mammalian cells. These findings suggest a link between cultivation at 32˚C, production of MLV-A and the Shh signalling pathway.

The promoter activity of the human c-fos and human cytomegalovirus (CMV) immediate early promoter was compared in transient and stable transfection experiments with six cell lines of mouse, human, and hamster origin which are all of commercial importance. The c-fos promoter was 1.8-5.6-fold stronger than the CMV promoter in BHK-A, BHK-B, CHO-DHFR(-), and mouse NIH-3T3 in stable transfectants and less effective in mouse myeloma or human 293 cells, suggesting a new transcriptional control element for high-level expression and protein production in mammalian cells. The induction profiles determined in the presence and absence of serum are dependent on the cell line used. Induction levels of up to 8-fold could be achieved in preselected cell pools.

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