Abundance of the primary transcript and its processed product of growth-related genes in normal and leukemic cells during proliferation and differentiation.
The relative abundance of primary transcript and mature mRNA of the c-myc, calcyclin, S14 ribosomal protein, and rRNA genes was determined densitometrically after reverse transcriptase-polymerase chain reaction and Northern blotting analysis in resting and mitogen-stimulated lymphocytes, proliferating and terminally differentiated HL-60 cells, and leukemic blast cells. Transcription and processing of c-myc and rRNA gene transcripts increased proportionally after mitogen stimulation, whereas these processes were independent of cell cycling status in the case of the S14 gene. Normal lymphocytes showed an unexpectedly large amount of primary transcript of the calcyclin gene, whereas the corresponding mRNA was undetectable. The abundance of c-myc, calcyclin, and S14 mRNA in terminally differentiated HL-60 cells decreased sharply, compared to their proliferating counterparts. This decrease reflected post-transcriptional modulation, since the abundance of precursor remained essentially unchanged. After HL-60 differentiation, the 32S rRNA levels remained relatively high, whereas the 45S primary transcript almost disappeared. Leukemic blast cells displayed highly variable precursor/mRNA ratios of c-myc, calcyclin, and S14 genes but consistently high ratios of 32S to 45S RNA, suggesting that the cleavage rate of the 32S rRNA was sharply reduced in these cells, resulting in an accumulation of this molecule. These results suggest the importance of efficient processing of primary transcript to generate adequate functional mRNA, thus regulating gene expression. Furthermore, in terminally differentiated cells and leukemic blast cells a stabilization of the primary transcript without efficient processing can be observed. The role of the stabilization of the primary transcript in terminal differentiation is further supported by the results of run-off transcription, indicating a sharp decrease of c-myc and calcyclin transcription rate in retinoic acid/dimethyl sulfoxide-treated HL-60 cells.
Ferrari
S
S
II Medical Clinic, University of Modena, Italy.
Tagliafico
E
E
Manfredini
R
R
Grande
A
A
Rossi
E
E
Zucchini
P
P
Torelli
G
G
Torelli
U
U
D000208:Acute Disease; D001483:Base Sequence; D015152:Blotting, Northern; D002135:Calcium-Binding Proteins / Q000235:genetics*; D018797:Cell Cycle Proteins; D002454:Cell Differentiation; D002455:Cell Division; D015342:DNA Probes; D015973:Gene Expression Regulation, Leukemic / Q000235:genetics*; D006801:Humans; D007951:Leukemia, Myeloid / Q000235:genetics; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction; D016271:Proto-Oncogene Proteins c-myc / Q000235:genetics*; D012333:RNA, Messenger / Q000032:analysis* / Q000235:genetics; D012269:Ribosomal Proteins / Q000235:genetics*; D000075383:S100 Calcium Binding Protein A6; D009418:S100 Proteins
D002135:Calcium-Binding Proteins; D018797:Cell Cycle Proteins; D015342:DNA Probes; D016271:Proto-Oncogene Proteins c-myc; D012333:RNA, Messenger; D012269:Ribosomal Proteins; D000075383:S100 Calcium Binding Protein A6; D009418:S100 Proteins; C039747:ribosomal protein S14; C055351:S100A6 protein, human