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The incorporation of tritiated thymidine, uridine and leucine, into the acid-precipitable material of DNA. RNA and proteins, respectively, was studied by autoradiography throughout the cell cycle of Amoeba proteus. DNA synthesis occupied the first 17 h of the cycle (57 h long) and 2 peaks between 0.5 and 9.13 h accounted for the majority of the thymidine incorporation. RNA synthesis was represented by a series of peak uridine grain counts, the 3 major peaks occurring at 10, 26-27 and 47-48 h. The incorporation of leucine also followed a pattern of peaks and dips, the main peaks occurring 1-2 h after the major increases in uridine incorporation. The fraction of label present over the nucleus decreased during the cell cycle, and this was probably due to a lowered incorporation of the leucine label by proteins synthesized in the cytoplasm and destined to become nuclear proteins. The incorporation patterns of 6 amino acids (arginine, aspartic acid, leucine, lysine, serine and valine) were studied individually during 3 periods of the cell cycle: 0-10 h (S phase); 20-30 h (early G2); and 40-50 h (mid-late G2). Variations in the intensity and timings of the incorporation maxima of the amino acids were observed, although the timings of increased grain counts of some of the amino acids frequently coincided. "Unique" incorporation peaks (i.e. only observed in one of the amino acids studied) possibly indicate the synthesis of phase-specific proteins. The amino acid and nucleoside incorporation profiles presented in this paper will enable the results obtained from future studies on amoebae to be related to the macromolecular synthesis patterns.

Recombinant plasmid cDNA libraries representing the polyadenylated RNAs in the myeloid cell lines KG1 and ML1 have been constructed. The screening protocol has identified several clones which contain sequences homologous to RNAs expressed at high abundance in one or more of the myeloid cell lines KG1, ML1 and HL60. The relative abundances of RNAs homologous to three recombinants, pML15, pKG21 and pKGA/F5 were measured by an RNA dot hybridisation method in total RNAs isolated from peripheral blood leukocytes from leukaemic patients and normal individuals. High levels of these RNAs were observed mainly in ANLL and acute phase CML samples. The data suggest that these probes have the potential to sub-divide the ANLLs and to extend a molecular classification of the myeloid leukaemias.

The relative concentrations of pCG14 RNA (a myelocyte-specific mRNA), pAM6 RNA (a monocyte-lineage specific marker), and c-myc RNA (present at higher concentrations in more primitive cells) were measured in the RNAs from peripheral blood leucocytes from leukaemic samples and normal individuals. The potential of differences in the relative abundances of these three RNAs in a series of 34 leukaemias was assessed as a means of distinguishing among the myeloid leukaemias. The chronic phase CGL samples were clustered with a high pCG14 RNA, a medium to low c-myc RNA abundance, and a variable pAM6 RNA level. The ANLL samples could be distinguished from the chronic phase CGL by virtue of different relative abundances of these RNAs: low pCG14, medium to high c-myc and a variable pAM6. The acute phase CGL samples showed a variety of relative RNA abundances with some samples sited within the ANLL region. Using samples obtained during the progression of CGL we have shown a shift in the relative abundances of these RNAs from the CGL region towards the ANLL region, and have suggested that the use of these parameters may allow the progression to acute phase to be monitored and, possibly, predicted.

Autoradiography has been used to examine the migration of proteins from a radioactively labelled amoeba nucleus following transplantation into an unlabelled homophasic amoeba. Nuclei were transferred at three times in the cell cycle coinciding with DNA synthesis (4 h post-division); a peak of RNA synthesis (25 h); and a relative lull in synthetic activity (43 h). Six amino acids were added individually to the culture medium to label the nuclear proteins. Migration of the proteins from the donor nucleus was found to be greatest following the transfer of [3H]aspartic acid-labelled nuclei and least with protein labelled with the basic amino acids. All amino acids exhibited the greatest extent of migration following the 25-h transfers, i.e., coinciding with a peak of RNA synthesis at 26-27.5 h. Actinomycin D (actD) inhibition of RNA synthesis reduced, but did not eliminate the extent of protein migration from the transplanted nucleus, thus indicating the existence of two classes of migratory proteins. Firstly, proteins, associated with RNA transport, which migrated mainly into the host cytoplasm. The second class migrated into the host nucleus from the transplanted nucleus, irrespective of RNa synthesis. The shuttling character of the latter class of proteins is consistent with a role of regulation of nuclear activity.