RNA-directed actions of 8-chloro-adenosine in multiple myeloma cells.
The purine analogue, 8-chloro-adenosine (8-Cl-Ado), induces apoptosis in a number of multiple myeloma (MM) cell lines. This ribonucleoside analogue accumulates as a triphosphate and selectively inhibits RNA synthesis without perturbing DNA synthesis. Cellular RNA is synthesized by one of three polymerases (Pol I, II, or III); thus, the inhibition of one or more RNA polymerases may be mediating 8-Cl-Ado cytotoxicity. Here, we have addressed this question by dissecting the RNA-directed actions of 8-Cl-Ado in MM cells. Differential alterations in [(3)H]uridine incorporation were found in the three major classes of RNA after a 20-h exposure with 10 microM 8-Cl-Ado. The synthesis rate of Pol III transcripts, 5 S and tRNA, remained unchanged, whereas Pol I-mediated rRNA synthesis decreased by approximately 20%. In contrast, mRNA synthesis, which is transcribed by Pol II, rapidly declined within 4 h and reached a 50% decrease, which was maintained for 20 h. Parallel to RNA synthesis inhibition, 8-Cl-Ado was maximally incorporated in the mRNA (>13 nmol/mg RNA), which was 5-fold higher than the tRNA and rRNA incorporation. Electrophoretic and radiographic analysis of newly synthesized and processed [(14)C]uridine-labeled transcripts indicated that the analogue blocks transcription elongation. Consistent with that result, high-performance liquid chromatography analysis of micrococcal nuclease and spleen phosphodiesterase-digested RNA demonstrated that the analogue incorporation is at the 3' terminus. In conclusion, our data demonstrate that in MM cells, 8-Cl-Ado is preferentially incorporated into mRNA, suggesting a propensity toward Pol II, and inhibits RNA synthesis by premature transcriptional chain termination.
Stellrecht
Christine M
CM
Department of Experimental Therapeutics, University of Texas M. D. Anderson Cancer Center, 1515 Holcombe Boulevard, Houston, TX 77030, USA.
Rodriguez
Carlos O
CO
Ayres
Mary
M
Gandhi
Varsha
V
D015762:2-Chloroadenosine / Q000031:analogs & derivatives* / Q000493:pharmacokinetics / Q000494:pharmacology*; D000255:Adenosine Triphosphate / Q000378:metabolism; D006801:Humans; D009101:Multiple Myeloma / Q000188:drug therapy* / Q000235:genetics / Q000378:metabolism*; D012333:RNA, Messenger / Q000096:biosynthesis / Q000378:metabolism; D012334:RNA, Neoplasm / Q000096:biosynthesis* / Q000378:metabolism; D012335:RNA, Ribosomal / Q000096:biosynthesis / Q000378:metabolism; D013379:Substrate Specificity; D014158:Transcription, Genetic / Q000187:drug effects
D012333:RNA, Messenger; D012334:RNA, Neoplasm; D012335:RNA, Ribosomal; D015762:2-Chloroadenosine; D000255:Adenosine Triphosphate; C075440:8-chloroadenosine