10. D N A A N D R N A S Y N T H E S I S
327
become radioactive, indicating that the transfer of activity occurred only
from nucleus to cytoplasm and not vice versa.
Many recent experiments with cell cultures of various types have
utilized tritiated cytidine as an R N A precursor. Although cytidine
is incorporated into both nucleic acids in rapidly-dividing cells, the
uptake into D N A is often only about 10°/o of that into R N A when
averaged over the cell population. Using an autoradiographic method
in conjunction with tritiated cytidine, Goldstein and Micou (1959a)
cultured a continuously propagated strain of cells originally derived
from human amnion. The autoradiographs showed that exposure of
the cells to tritiated cytidine for 1 h resulted in nuclei only being labelled;
however, if the cells were placed in medium containing excess unlabelled
cytidine for several hours after the initial incubation in radioactive
precursor, the cytoplasm also became labelled. Extraction with RNAase
showed that part of the nuclear activity and all the cytoplasmic activity
was incorporated in the R N A . In additional experiments (Goldstein
and Micou, 1959b), very little uptake was observed in the nucleus after
2 min incubation in tritiated cytidine, but after 5 min incubation both
chromatin and nucleolus were labelled and after 10 min there was an
increase of nucleolar label with respect to chromatin label. This result
could mean merely that different rates of incorporation of precursor
occur in R N A synthesized in the nucleolus and chromatin respectively;
on the other hand, it has been suggested (Woods, 1960) that R N A
synthesized in the chromatin is transferred to the nucleolus (see also
the review by Sirlin, 1962). Both Goldstein and Micou (1959a, b) and
Woods (1960) interpret their results to mean that some R N A is synthesized in both nucleolus and chromatin and is later transferred to the
cytoplasm.
Substantially similar results for the kinetics of cytidine incorporation
were obtained in HeLa cells by Feinendegen, Bond, Shreeve and Painter
(1960). Autoradiographic grain-counts showed that the most rapid
uptake was initially in the chromatin, accompanied by a somewhat
slower uptake by the nucleolus. After a further hour in medium containing excess unlabelled cytidine, the label began to appear in the
cytoplasm. In other experiments, it was demonstrated that after
removal of the tritiated cytidine from the medium, tritium label
continued to be incorporated into both R N A and D N A for some time,
so that after a 6 h incubation only 40°/o of the cell population had D N A
labelling, but after a further 16 h in non-radioactive medium almost
all the cells contained labelled D N A . This label, which continued to
be incorporated, could not entirely be diluted out of the medium by
addition of excess unlabelled cytidine, and Feinendegen and his
collaborators proposed that, although a major part of the R N A is
327
become radioactive, indicating that the transfer of activity occurred only
from nucleus to cytoplasm and not vice versa.
Many recent experiments with cell cultures of various types have
utilized tritiated cytidine as an R N A precursor. Although cytidine
is incorporated into both nucleic acids in rapidly-dividing cells, the
uptake into D N A is often only about 10°/o of that into R N A when
averaged over the cell population. Using an autoradiographic method
in conjunction with tritiated cytidine, Goldstein and Micou (1959a)
cultured a continuously propagated strain of cells originally derived
from human amnion. The autoradiographs showed that exposure of
the cells to tritiated cytidine for 1 h resulted in nuclei only being labelled;
however, if the cells were placed in medium containing excess unlabelled
cytidine for several hours after the initial incubation in radioactive
precursor, the cytoplasm also became labelled. Extraction with RNAase
showed that part of the nuclear activity and all the cytoplasmic activity
was incorporated in the R N A . In additional experiments (Goldstein
and Micou, 1959b), very little uptake was observed in the nucleus after
2 min incubation in tritiated cytidine, but after 5 min incubation both
chromatin and nucleolus were labelled and after 10 min there was an
increase of nucleolar label with respect to chromatin label. This result
could mean merely that different rates of incorporation of precursor
occur in R N A synthesized in the nucleolus and chromatin respectively;
on the other hand, it has been suggested (Woods, 1960) that R N A
synthesized in the chromatin is transferred to the nucleolus (see also
the review by Sirlin, 1962). Both Goldstein and Micou (1959a, b) and
Woods (1960) interpret their results to mean that some R N A is synthesized in both nucleolus and chromatin and is later transferred to the
cytoplasm.
Substantially similar results for the kinetics of cytidine incorporation
were obtained in HeLa cells by Feinendegen, Bond, Shreeve and Painter
(1960). Autoradiographic grain-counts showed that the most rapid
uptake was initially in the chromatin, accompanied by a somewhat
slower uptake by the nucleolus. After a further hour in medium containing excess unlabelled cytidine, the label began to appear in the
cytoplasm. In other experiments, it was demonstrated that after
removal of the tritiated cytidine from the medium, tritium label
continued to be incorporated into both R N A and D N A for some time,
so that after a 6 h incubation only 40°/o of the cell population had D N A
labelling, but after a further 16 h in non-radioactive medium almost
all the cells contained labelled D N A . This label, which continued to
be incorporated, could not entirely be diluted out of the medium by
addition of excess unlabelled cytidine, and Feinendegen and his
collaborators proposed that, although a major part of the R N A is