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The model of RNA macromolecular structure
TIBS 1 5 - J U N E 1 9 9 0
LETTERS The modelof RNA macromolecularstructure In his recent interesting historical essay on RNA spatial organization, Bogdanov ~ attempts to correct what he believes to be an omission in textbooks and historical treatises of the creation of the fundamental model of RNA macromolecular structure in the late 1950s. He is indebted to Drs H. Boedtker, P. Doty and A. S. Spirin for allowing him to test their memories concerning their seminal contributions relating to the structure of tobacco mosaic virus (TMV) RNA. We have also tested our memories and recall that we independently developed procedures for isolating highly polymerized RNA from E. coil protoplasts, showed this ribosomal RNA to be composed of three boundaries z, and demonstrated by viscosity and streaming birefringence measurements over a wide range of salt concentrations and potentiometric titration that, unlike DNA, RNA behaved as a typical contractile single-stranded polyelectrolyte chain 3. These observations were then reported by us in detail in 19594. To give a historical flavor to these
Substrate specificityand the mdr pump I would like to comment on the recent thought-provoking article by Ian West I on the substrate specificity of the multi-drugresistance (mdr) pump. In it he suggests that specificity may be achieved by tagging drugs with a glutathione group. As a former worker in the multi-drugresistance field it has never struck me as odd that the P-glycoprotein should know which chemicals to extrude from the cell. The experimental model systems are tumour cell lines which have been made highly drug-resistant in vitro by selecting for overproduction of the pump in response to the presence of a cytotoxic drug. I believe that West confuses this rather abnormal state of affairs, which the average somatic cell is unlikely ever to encounter, with the normal situation of a kidney or liver cell expressing the mdrl gene. The normal function of the mdrf pump and the spectrum of its substrates are not known but one can make an educated guess. The elevated levels of glutathione-S-transferase (GST) observed
218
exciting early findings we refer to an English translation of excerpts of a letter in Hebrew sent by UL to HE (at that time at the Mellon Institute in Pittsburgh) after the 4th International Congress of Biochemistry in Vienna, dated 14 October 19585. We reported a similar high molecular-weight structure for rat liver RNA6 and soon thereafter the secondary folding of the E. coil RNA7 and tRNAa. The reports referred to above were given ample mention in the 1960 review by Spirin a as well as in the review by Doty the same year ~°. Lest these remarks sound as motivated only towards the maintenance of a correct historical record, we would like to recall here an observation connecting the TMV and E. coil work. We had noticed 4 that the positive birefringence of flow observed by us for E. coli RNA matched that reported by Rosalind Franklin in TMV suspensions 11. Whereas birefringence of E. coil RNA disappears upon addition of salt (0.2 M NaCI), and that of TMV disappears upon removal of the RNA constituents from the virus particles, in solutions of DNA considerable negative birefringence of flow persists at neutral pH, even in the highest concentrations of NaCI (2 M) that
in some resistant cell lines may simply be the fortuitous result of a common pathway regulating cellular detoxification. To my knowledge, there is no evidence that GST levels rise in proportion to the P-glycoprotein levels in highly drugresistant cell lines. West notes that there is a correlation between the sites of glutathione-conjugate excretion and of mdrl expression in certain normal cell types, but such a correlation does not extend to cell types that can become cancerous and subsequently drugresistant. For the latter an active mdrl gene is not required. It can be activated under drug selection. I also do not agree that mdrl might be a gene family. Extensive genomic mapping and hybridization studies by myself and others 2,3 leave room for at most three mdr genes. Of these mdrl is both necessary and sufficient to generate the multi-drug resistance phenotype characterized by a variable crossresistance pattern to drugs to which the cell may not have been exposed before 4. The above observations together with those highlighted by West render a role for GST in multi-drug resistance improbable. This leaves us
had been investigated. This gave a clear indication of the considerable difference between DNA and RNA structure.
References I Bogdanov,A. A. (1989) Trends Biochem. Sci. 14, 505-507 2 Littauer, U. Z. and Eisenberg, H. (1958) Bull. Res. Counc. Israel 7A, 114-115 3 Eisenberg, H. and Littauer, U. Z. (1958) Bull. Res. Counc. Israel 7A, 115 4 Littauer, U. Z. and Eisenberg, H. (1959) Biochim. Biophys. Acta 32, 320-337 5 Eisenberg, H. in Comprehensive Biochemistry, Vol. 37 (Semenza,G. and Jaenicke, R., eds) Elsevier (in press) 6 Laskow, R., Margoliash, E., Littauer, U. Z. and Eisenberg, H. (1959) Biochim. Biophys. Acta 33, 247-248 7 Cox, R. A. and Littauer, U. Z. (1959) Nature 184, 818-819 8 Littauer, U. Z. (1961)in Protein Biosynthesis (Harris, R. J. C., ed.), pp. 143-162, Academic Press 9 Spirin, A. S. (1960) J. Mol. Biol. 2, 436-446 10 Doty, P. (1961) The Harvey Lectures 55, 103-139 11 Franklin, R. E. (1955) Biochim. Biophys. Acta 18, 313-314
U. Z. LITrAUER AND H. EISENBERG The Weizmann Institute of Science, Rehovot 76100, Israel.
with just one mdr pump, and the problem of substrate specificity. The variable crossresistance pattern displayed by multi-drug-resistant cells must be the result of the mdrl pump not being able to extrude all drugs with the same efficiency. This by itself precludes a role for uniform chemical tagging in conferring substrate specificity. An explanation 5 which, though untested, is both simple and attractive might be that the mdrl pump has a broad specificity for compounds that perhaps are hydrophobic, aromatic and/or possess a carbonyl group but also has a broad spectrum of affinities for the compounds individually (the affinity for compounds such as tryptophan and guanine is assumed to be nil). It could explain the curious findings of Cornwell et al. 6, Safa et al. 7 and Horio et al. s that puromycin, actinomycin D and colchicine fail to compete effectively with vinblastin for binding to the mdrl pump, even though in one case 6 overexpression of the pump was selected for by exposure to colchicine. Apparently the pump has a higher affinity for vinblastin than for the other three drugs but comparable affinity
LETTERS The modelof RNA macromolecularstructure In his recent interesting historical essay on RNA spatial organization, Bogdanov ~ attempts to correct what he believes to be an omission in textbooks and historical treatises of the creation of the fundamental model of RNA macromolecular structure in the late 1950s. He is indebted to Drs H. Boedtker, P. Doty and A. S. Spirin for allowing him to test their memories concerning their seminal contributions relating to the structure of tobacco mosaic virus (TMV) RNA. We have also tested our memories and recall that we independently developed procedures for isolating highly polymerized RNA from E. coil protoplasts, showed this ribosomal RNA to be composed of three boundaries z, and demonstrated by viscosity and streaming birefringence measurements over a wide range of salt concentrations and potentiometric titration that, unlike DNA, RNA behaved as a typical contractile single-stranded polyelectrolyte chain 3. These observations were then reported by us in detail in 19594. To give a historical flavor to these
Substrate specificityand the mdr pump I would like to comment on the recent thought-provoking article by Ian West I on the substrate specificity of the multi-drugresistance (mdr) pump. In it he suggests that specificity may be achieved by tagging drugs with a glutathione group. As a former worker in the multi-drugresistance field it has never struck me as odd that the P-glycoprotein should know which chemicals to extrude from the cell. The experimental model systems are tumour cell lines which have been made highly drug-resistant in vitro by selecting for overproduction of the pump in response to the presence of a cytotoxic drug. I believe that West confuses this rather abnormal state of affairs, which the average somatic cell is unlikely ever to encounter, with the normal situation of a kidney or liver cell expressing the mdrl gene. The normal function of the mdrf pump and the spectrum of its substrates are not known but one can make an educated guess. The elevated levels of glutathione-S-transferase (GST) observed
218
exciting early findings we refer to an English translation of excerpts of a letter in Hebrew sent by UL to HE (at that time at the Mellon Institute in Pittsburgh) after the 4th International Congress of Biochemistry in Vienna, dated 14 October 19585. We reported a similar high molecular-weight structure for rat liver RNA6 and soon thereafter the secondary folding of the E. coil RNA7 and tRNAa. The reports referred to above were given ample mention in the 1960 review by Spirin a as well as in the review by Doty the same year ~°. Lest these remarks sound as motivated only towards the maintenance of a correct historical record, we would like to recall here an observation connecting the TMV and E. coil work. We had noticed 4 that the positive birefringence of flow observed by us for E. coli RNA matched that reported by Rosalind Franklin in TMV suspensions 11. Whereas birefringence of E. coil RNA disappears upon addition of salt (0.2 M NaCI), and that of TMV disappears upon removal of the RNA constituents from the virus particles, in solutions of DNA considerable negative birefringence of flow persists at neutral pH, even in the highest concentrations of NaCI (2 M) that
in some resistant cell lines may simply be the fortuitous result of a common pathway regulating cellular detoxification. To my knowledge, there is no evidence that GST levels rise in proportion to the P-glycoprotein levels in highly drugresistant cell lines. West notes that there is a correlation between the sites of glutathione-conjugate excretion and of mdrl expression in certain normal cell types, but such a correlation does not extend to cell types that can become cancerous and subsequently drugresistant. For the latter an active mdrl gene is not required. It can be activated under drug selection. I also do not agree that mdrl might be a gene family. Extensive genomic mapping and hybridization studies by myself and others 2,3 leave room for at most three mdr genes. Of these mdrl is both necessary and sufficient to generate the multi-drug resistance phenotype characterized by a variable crossresistance pattern to drugs to which the cell may not have been exposed before 4. The above observations together with those highlighted by West render a role for GST in multi-drug resistance improbable. This leaves us
had been investigated. This gave a clear indication of the considerable difference between DNA and RNA structure.
References I Bogdanov,A. A. (1989) Trends Biochem. Sci. 14, 505-507 2 Littauer, U. Z. and Eisenberg, H. (1958) Bull. Res. Counc. Israel 7A, 114-115 3 Eisenberg, H. and Littauer, U. Z. (1958) Bull. Res. Counc. Israel 7A, 115 4 Littauer, U. Z. and Eisenberg, H. (1959) Biochim. Biophys. Acta 32, 320-337 5 Eisenberg, H. in Comprehensive Biochemistry, Vol. 37 (Semenza,G. and Jaenicke, R., eds) Elsevier (in press) 6 Laskow, R., Margoliash, E., Littauer, U. Z. and Eisenberg, H. (1959) Biochim. Biophys. Acta 33, 247-248 7 Cox, R. A. and Littauer, U. Z. (1959) Nature 184, 818-819 8 Littauer, U. Z. (1961)in Protein Biosynthesis (Harris, R. J. C., ed.), pp. 143-162, Academic Press 9 Spirin, A. S. (1960) J. Mol. Biol. 2, 436-446 10 Doty, P. (1961) The Harvey Lectures 55, 103-139 11 Franklin, R. E. (1955) Biochim. Biophys. Acta 18, 313-314
U. Z. LITrAUER AND H. EISENBERG The Weizmann Institute of Science, Rehovot 76100, Israel.
with just one mdr pump, and the problem of substrate specificity. The variable crossresistance pattern displayed by multi-drug-resistant cells must be the result of the mdrl pump not being able to extrude all drugs with the same efficiency. This by itself precludes a role for uniform chemical tagging in conferring substrate specificity. An explanation 5 which, though untested, is both simple and attractive might be that the mdrl pump has a broad specificity for compounds that perhaps are hydrophobic, aromatic and/or possess a carbonyl group but also has a broad spectrum of affinities for the compounds individually (the affinity for compounds such as tryptophan and guanine is assumed to be nil). It could explain the curious findings of Cornwell et al. 6, Safa et al. 7 and Horio et al. s that puromycin, actinomycin D and colchicine fail to compete effectively with vinblastin for binding to the mdrl pump, even though in one case 6 overexpression of the pump was selected for by exposure to colchicine. Apparently the pump has a higher affinity for vinblastin than for the other three drugs but comparable affinity