Glycoprotein synthesis in a temperature‐sensitive Chinese hamster cell cycle mutant

Publisher Website
Google Scholar
Abstract
A temperature‐sensitive mutant of Chinese hamster cells is described which has two interesting properties: (1) it is a cell cycle mutant and (2) glycoprotein synthesis appears to be affected at the non‐permissive temperature (40°C). Synchronized cells shifted to 40°C in the beginning of their G1 phase do not incorporate [3H]‐thymidine into DNA during the expected S‐phase, but once DNA synthesis has been initiated (∼ 10 hours after termination of serum starvation) a shift to 40°C no longer leads to an arrest of DNA synthesis. Flow microfluorimetric analysis of DNA content/cell supports this conclusion and indicates that a majority of cells become arrested in the G1 phase of the cell cycle when a non‐synchronized population of cells is transferred to 40°C. Apparently at all times in the cell cycle there is a drastic reduction of incorporation of labeled sugars (particularly fucose) into glycoproteins. The uptake of fucose and its conversion to GDP‐fucose appears to be normal at 40°C. Chromatographic analysis indicates that all classes of glycoproteins are affected, and we do not find any evidence for partially completed oligosaccharides at 40°C. Overall protein synthesis is not reduced at the nonpermissive temperature during the time interval under consideration and the number of polysomes attached to membranes (RER) is also normal at 40°C. This suggests that the defect is at an early step in the synthesis or regulation of synthesis of glycoproteins. The mutation is a recessive mutation in hybrid cells and mutagen induced revertants can be obtained which grow normally at 40°C and in which glycoprotein synthesis at 40°C is restored to normal, wild type levels.