Poly(A) Tail Shortening by a Mammalian Poly(A)-specific 3′-Exoribonuclease

Abstract

3′-Exonucleolytic removal of the poly(A) tail is the first and often rate-limiting step in the decay of many eucaryotic mRNAs. In a cytoplasmic extract from HeLa cells, the poly(A) tail of mRNA was degraded from the 3′-end. In agreement with earlierin vivo observations, prominent decay intermediates differed in length by about 30 nucleotides. The Mg2+-dependent, poly(A)-specific 3′-exoribonuclease responsible for this poly(A) shortening activity was purified from calf thymus. A polypeptide of 74 kDa copurified with the activity. The deadenylating nuclease (DAN) required a free 3′-OH group, released solely 5′-AMP, degraded RNA in a distributive fashion, and preferred poly(A) as a substrate. At low salt concentration, the activity of purified DAN was strongly dependent on spermidine or other, yet unidentified factors. Under these reaction conditions, DAN was also stimulated by the cytoplasmic poly(A)-binding protein I (PAB I). At physiological salt concentration, the stimulatory effect of spermidine was weak and PAB I was inhibitory. At either salt concentration DAN and PAB I reconstituted poly(A) shortening with the same pattern of intermediates seen in cytoplasmic extract. The properties of DAN suggest that the enzyme might be involved in the deadenylation of mRNA in vivo. 3′-Exonucleolytic removal of the poly(A) tail is the first and often rate-limiting step in the decay of many eucaryotic mRNAs. In a cytoplasmic extract from HeLa cells, the poly(A) tail of mRNA was degraded from the 3′-end. In agreement with earlierin vivo observations, prominent decay intermediates differed in length by about 30 nucleotides. The Mg2+-dependent, poly(A)-specific 3′-exoribonuclease responsible for this poly(A) shortening activity was purified from calf thymus. A polypeptide of 74 kDa copurified with the activity. The deadenylating nuclease (DAN) required a free 3′-OH group, released solely 5′-AMP, degraded RNA in a distributive fashion, and preferred poly(A) as a substrate. At low salt concentration, the activity of purified DAN was strongly dependent on spermidine or other, yet unidentified factors. Under these reaction conditions, DAN was also stimulated by the cytoplasmic poly(A)-binding protein I (PAB I). At physiological salt concentration, the stimulatory effect of spermidine was weak and PAB I was inhibitory. At either salt concentration DAN and PAB I reconstituted poly(A) shortening with the same pattern of intermediates seen in cytoplasmic extract. The properties of DAN suggest that the enzyme might be involved in the deadenylation of mRNA in vivo. Gene expression depends on the concentration of mRNA, which in turn is determined by the ratio of transcription and turnover. Each mRNA has its own specific half-life, which can sometimes be altered in response to internal or external signals. Rapid degradation is particularly important for mRNAs encoding proteins that are only transiently required, like those of the early response to serum stimulation (1Ross J. Microbiol. Rev. 1995; 59: 423-450Crossref PubMed Google Scholar, 2Schiavi S.C. Belasco J.G. Greenberg M.E. Biochim. Biophys. Acta. 1992; 1114: 95-106Crossref PubMed Scopus (94) Google Scholar). Whereas it is obvious that mRNA instability is essential for a quick shut-off of protein synthesis after transient transcription, a closer look reveals that a short mRNA half-life also favors rapid induction (1Ross J. Microbiol. Rev. 1995; 59: 423-450Crossref PubMed Google Scholar). In yeast, a pathway of decay has been described that is common to many mRNAs, both stable and unstable (3Muhlrad D. Decker C.J. Parker R. Mol. Cell. Biol. 1995; 15: 2145-2156Crossref PubMed Scopus (263) Google Scholar, 4Muhlrad D. Decker C.J. Parker R. Genes Dev. 1994; 8: 855-866Crossref PubMed Scopus (412) Google Scholar, 5Caponigro G. Parker R. Microbiol. Rev. 1996; 60: 233-249Crossref PubMed Google Scholar). Degradation is initiated by shortening of the poly(A) tail. Only after the poly(A) tail has been shortened to 15 nucleotides or less is the 5′-cap removed. Cap removal exposes the mRNA to a 5′-3′-exonuclease. Certain sequences within the RNA determine the rate of poly(A) shortening (6Decker C.J. Parker R. Genes Dev. 1993; 7: 1632-1643Crossref PubMed Scopus (527) Google Scholar, 7Muhlrad D. Parker R. Genes Dev. 1992; 6: 2100-2111Crossref PubMed Scopus (160) Google Scholar). Mutations in these sequences lead to altered rates of deadenylation and often to corresponding changes in mRNA half-life (5Caponigro G. Parker R. Microbiol. Rev. 1996; 60: 233-249Crossref PubMed Google Scholar). Thus, deadenylation is not only the first step in mRNA decay, but can be rate-limiting (5Caponigro G. Parker R. Microbiol. Rev. 1996; 60: 233-249Crossref PubMed Google Scholar, 8Jacobson A. Peltz S.W. Annu. Rev. Biochem. 1996; 65: 693-739Crossref PubMed Scopus (577) Google Scholar). In yeast, poly(A) tail removal requires the cytoplasmic poly(A)-binding protein I (PAB I). 1The abbreviations used are: PAB I, poly(A)-binding protein I; PAB II, poly(A)-binding protein II; DAN, deadenylating nuclease; PAN, poly(A) nuclease. In strains lacking PAB I, the steady state population of mRNA has longer poly(A) tails compared with wild type cells (9Sachs A.B. Davis R.W. Cell. 1989; 58: 857-867Abstract Full Text PDF PubMed Scopus (392) Google Scholar). The PGK1 and MFA2 mRNAs are deadenylated only after a lag phase, and 3- and 6-fold more slowly, respectively, in these mutants (10Caponigro G. Parker R. Genes Dev. 1995; 9: 2421-2432Crossref PubMed Scopus (232) Google Scholar). Extracts prepared from Δpab1 strains do not degrade poly(A) (11Sachs A.B. Deardorff J.A. Cell. 1992; 70: 961-973Abstract Full Text PDF PubMed Scopus (160) Google Scholar). Addition of recombinant PAB I restores deadenylation. A Mg2+-dependent poly(A)-specific 3′-exonuclease (poly(A) nuclease (PAN)) requiring PAB I for its activity has been purified. The enzyme contains two polypeptides of 76 and 127 kDa (11Sachs A.B. Deardorff J.A. Cell. 1992; 70: 961-973Abstract Full Text PDF PubMed Scopus (160) Google Scholar, 12Lowell J.E. Rudner D.Z. Sachs A.B. Genes Dev. 1992; 6: 2088-2099Crossref PubMed Scopus (91) Google Scholar, 13Boeck R. Tarun Jr., S. Rieger M. Deardorff J.A. Müller-Auer S. Sachs A.B. J. Biol. Chem. 1996; 271: 432-438Abstract Full Text Full Text PDF PubMed Scopus (147) Google Scholar, 14Brown C.E. Tarun S.Z. Boeck R. Sachs A.B. Mol. Cell. Biol. 1996; 16: 5744-5753Crossref PubMed Scopus (130) Google Scholar). At low salt concentration, PAN does not attack naked poly(A). The requirement for a RNA-binding cofactor can be satisfied not only by PAB I but also by spermidine (12Lowell J.E. Rudner D.Z. Sachs A.B. Genes Dev. 1992; 6: 2088-2099Crossref PubMed Scopus (91) Google Scholar). At high salt concentration, PAN has a PAB I- and spermidine-independent activity. It is not yet clear whether PAN is the enzyme responsible for deadenylation in vivo (14Brown C.E. Tarun S.Z. Boeck R. Sachs A.B. Mol. Cell. Biol. 1996; 16: 5744-5753Crossref PubMed Scopus (130) Google Scholar). Enzymes that catalyze decapping and 5′-exonucleolytic decay have also been purified, and genes encoding them have been cloned (5Caponigro G. Parker R. Microbiol. Rev. 1996; 60: 233-249Crossref PubMed Google Scholar, 8Jacobson A. Peltz S.W. Annu. Rev. Biochem. 1996; 65: 693-739Crossref PubMed Scopus (577) Google Scholar). In higher eucaryotes, the mechanism of mRNA decay is less clear. However, for many mRNAs it has been shown that poly(A) removal precedes the decay of the mRNA body (1Ross J. Microbiol. Rev. 1995; 59: 423-450Crossref PubMed Google Scholar, 2Schiavi S.C. Belasco J.G. Greenberg M.E. Biochim. Biophys. Acta. 1992; 1114: 95-106Crossref PubMed Scopus (94) Google Scholar, 15Wilson T. Treisman R. Nature. 1988; 336: 396-399Crossref PubMed Scopus (506) Google Scholar). As in yeast, mutations in the 3′-untranslated region or the coding region can change the rate of deadenylation and the mRNA half-life (8Jacobson A. Peltz S.W. Annu. Rev. Biochem. 1996; 65: 693-739Crossref PubMed Scopus (577) Google Scholar). While deadenylation is usually the prelude to degradation, this is not necessarily the case. In animal oocytes, deadenylation is used to stop the translation of certain mRNAs. The deadenylated RNAs are stored inactive but intact and may even be readenylated and used again at a later time (16Curtis J. Lehmann R. Zamore P.D. Cell. 1995; 81: 171-178Abstract Full Text PDF PubMed Scopus (368) Google Scholar, 17Wickens M. Trends Biochem. Sci. 1990; 15: 320-324Abstract Full Text PDF PubMed Scopus (114) Google Scholar). The enzymology of mRNA decay, and in particular of deadenylation, in higher eucaryotes is unknown. It is also uncertain which role PAB I plays in deadenylation. Depletion of PAB I from a cell extract promoted rapid deadenylation and decay of mRNA (18Bernstein P. Peltz S.W. Ross J. Mol. Cell. Biol. 1989; 9: 659-670Crossref PubMed Scopus (297) Google Scholar). Thus, it was suggested that, in contrast to the situation in yeast, PAB I in mammalian cells inhibits mRNA decay by protecting the poly(A) tail. However, since the experiments were carried out in vitro, it is not certain whether the nuclease degrading the poly(A) tail was the same that is responsible for this process in vivo. Overexpression of PAB I in Xenopus oocytes protected mRNA from deadenylation, again suggesting that PAB I may inhibit poly(A) removal rather it M. J. 1996; 15: PubMed Scopus Google Scholar). However, since deadenylation is used for the of in oocytes and is not by degradation of the it is not clear whether the deadenylation reaction in oocytes is the same as in responsible for poly(A) tail removal have not been in higher A poly(A)-specific 3′-exoribonuclease has been purified from HeLa cell extract J. A. A. J. PubMed Scopus Google Scholar, J. A. A. J. Biol. Chem. 1992; Full Text PDF PubMed Google Scholar). A 3′-exoribonuclease of kDa has been purified to nuclease is to degrade but it is not The enzyme is by PAB I Ross J. J. Biol. Chem. 1994; Full Text PDF PubMed Google Scholar). have a of poly(A) tail in vivo with prominent in length by nucleotides. a was described in the steady state population of mRNA PubMed Scopus Google Scholar, A. G. J. Biol. Chem. Full Text PDF PubMed Google Scholar, J.A. R. J. Biochem. PubMed Scopus Google less also in mRNAs for and M.E. S.W. 1996; PubMed Scopus Google Scholar, M. 1993; PubMed Scopus Google Scholar). were as deadenylation intermediates of the mRNA M. 1993; PubMed Scopus Google Scholar). A of intermediates of poly(A) tail shortening can also be seen in of mRNA T. Treisman R. Nature. 1988; 336: 396-399Crossref PubMed Scopus (506) Google the particular type of carried out of poly(A) tail PAB I about nucleotides to poly(A) J. Cell. Biol. PubMed Scopus Google Scholar). the the properties of PAB that, in a cytoplasmic extract of HeLa cells, the poly(A) tail is shortened from the 3′-end. The population of deadenylation intermediates has a length of about 30 nucleotides. have purified a poly(A)-specific 3′-exoribonuclease from calf thymus. of mRNA by the purified nuclease in the of PAB I in the same pattern of intermediates as seen in cytoplasmic and in vivo. and were with and J. T. A Scholar). of poly(A) was carried out as of poly(A) nucleotides was with of and of poly(A) for reaction for J. J. Biol. Chem. Full Text PDF PubMed Google Scholar). the of of about 30 nucleotides in were with of of and of poly(A) for poly(A) with either a 3′-OH or a of and was prepared as poly(A) was in for and at The were by the of of and were in The RNA was and was with the activity of in the by the were a and poly(A) the was RNAs were from the and contains the of the transcription which is of its poly(A) G. Cell. 1988; Full Text PDF PubMed Scopus Google Scholar). The is a a from the of the The at the in the coding region and at the in the 3′-untranslated region A. T. Cell. Full Text PDF PubMed Scopus Google Scholar). was from the by J. of by reaction as were of the and at the was and of was with and RNAs were by in transcription of the with RNA in the of and of as described J. Biol. Chem. 1996; 271: Full Text Full Text PDF PubMed Scopus (147) Google that the concentration was or and the concentration were prepared by the of poly(A) tail with poly(A) was used for of RNA at 30 for of J. J. Biol. Chem. Full Text PDF PubMed Google Scholar). the of poly(A) used for the RNA were as RNA was in a A the RNA was out and in of at The RNAs were with and PAB I was purified from calf as described A. P. P. J. Biol. Chem. 1993; Full Text PDF PubMed Google Scholar). A PAB I was a from M. and G. of recombinant poly(A) at the was a from P. of and HeLa cell cytoplasmic extract was by of extract was prepared from HeLa cell to and RNA A Scholar). of the the was with and and used as cytoplasmic extract. RNA and calf were from and serum were from was from nuclease from S. from and from was from and were from was the same as described S. J. Biol. Chem. Full Text PDF PubMed Google Scholar). was prepared to 65: PubMed Scopus Google Scholar, that was and were from degrading activity was by the of from poly(A) from 12Lowell J.E. Rudner D.Z. Sachs A.B. Genes Dev. 1992; 6: 2088-2099Crossref PubMed Scopus (91) Google Scholar). In used to the to of protein was at with of reaction serum of poly(A) and of poly(A) of The reaction was by the of the protein and at for 15 and by the of of The were at for 15 in a of the was to a of of and of The of nucleotides released was determined by was as the activity that of that the were in the of the the nuclease was that more of the was of the reaction for the purified the of reaction were spermidine with mRNA were at as of reaction lacking spermidine was with of or RNA and RNA were as PAB I or spermidine was to of or respectively, as and the reaction was at The reaction was by the of nuclease or HeLa cell cytoplasmic extract. At were with of and of of of and of for at 30 at the RNA was with in for at and on J. T. A Scholar). The were either to or with a The decay of poly(A) and was as of was to of reaction The reaction was by the of nuclease and at were with of for at and to a In of the decay of were with of were with and to a The were with of and nucleotides were In of the decay of poly(A) and was as of were to of reaction were by the of nuclease and at were with of and and on as described protein were by the of Biochem. PubMed Scopus Google Scholar). protein was determined by Nature. PubMed Scopus Google with serum as a of poly(A) decay were by as described Ross J. J. Biol. Chem. 1994; Full Text PDF PubMed Google Scholar). The was carried out at or to the were in and stored at The was and for to in this was prepared from of calf in as described J. Biol. Chem. Full Text PDF PubMed Google Scholar). The extract was to a which was with and with a from to at from to were to with and for on the was to with and again on and The was in of for of and by A was with and the was The was with of and with of at At this from two from of were and by with the was in of for of the same and was to a with The was with of and with a from to at DAN activity in a from to were and for 30 The was and was with The was with two and with a from to at DAN activity in a from both were for of 30 and to a with DAN was with at were to a with at of were to with and The was to a with The was with of of and with a from to at DAN activity in a cytoplasmic extract of HeLa cells, RNA a shortening of the poly(A) tail. decay intermediates differed in length by about 30 and the RNAs the reaction carried a poly(A) tail of about 15 nucleotides A The of deadenylated RNA that in these was but The same degradation pattern was at or and with of of the extract by step from a and the deadenylating activity was in the that this the same pattern of deadenylation intermediates as the extract. The step of the poly(A) degrading activity from the as determined by a that the of from poly(A) not A that the step also of the PAB I in the extract not on these and those described the decay pattern to the of a deadenylating nuclease and PAB extract was for the of the deadenylating activity. were for poly(A) degrading activity with a activity in a was to the poly(A) used as a substrate. Under these conditions, of a only a activity at corresponding to the activity in the step of the HeLa cell cytoplasmic extract. The poly(A) degrading activity be stimulated by spermidine A that the PAB I was to the nuclease not the the activity was with only poly(A) as a was The is in The on is of the nuclease with from the same for removal of a stimulatory The poly(A) degrading activity was purified about as the activity in the first the is A of the is shown in A. A prominent of about 74 kDa with the activity The same was also with the activity on a The poly(A) degrading activity was DAN, for deadenylating of the deadenylating was by the with was as described in a The was by the with was as described The DAN determined with the was at with spermidine and in the of At and the activity to 30 and The on was and not be by or In the of DAN activity was dependent on spermidine with of At this concentration of or was at activity was in the of The was with either or Under these conditions, the activity was at spermidine in the of At the activity be by or poly(A) was with DAN, and the decay were on a the were after DAN and shortened decay intermediates were In with poly(A) was in the even after but shortened intermediates were after with higher of DAN, poly(A) was degraded to not A of the same by that, with the nucleotides were released at and the reaction was within was released at and the the time of the not these that DAN is a released by DAN from poly(A) were by and were used as The released with As for a nuclease degrading poly(A) from the and 5′-AMP, DAN requires a free A with a was not degraded at of this with which has a poly(A) was degraded not a of DAN, a of the poly(A) was shortened in a of DAN or and the of the are on the that the is Thus, DAN after removal of or a nucleotides and and RNA A distributive of of DAN was also suggested by experiments in which a of poly(A) was with of The length of degradation intermediates with the of DAN A and not of of and were in of reaction with of DAN for at were on a of and are on of poly(A) or were in of reaction either spermidine and salt or and were with of DAN at were at the and as A of the was not degraded at was from the and with poly(A) a free 3′-OH for and reaction for The poly(A) was not poly(A). Thus, the of the poly(A) was and this was the for its to The DAN activity degraded poly(A) and but not and that of the used for this be either by poly(A) with in the of or by and in the of Thus, were not responsible for the of and was not DAN is a spermidine poly(A) or were degraded A and the was Under these reaction conditions, DAN degraded only poly(A) but not A that about of poly(A) been shortened after a was In the same was degraded in the of The of DAN was also with RNA as a the activity was the poly(A) tail was within The deadenylated body of the RNA and was degraded were with RNA as a that the deadenylated RNA stable even after not Thus, DAN is but not specific for poly(A). DAN was also specific for the poly(A) tail the spermidine-independent activity was at and mRNA deadenylation by A was and RNA in of reaction The RNA was at with or of DAN in the of or of PAB I or in its The ratio of PAB I is at the were at the and on a of are on the and decay are on the RNA was also used to the of PAB I on DAN activity. at spermidine PAB I were poly(A) tails were stable with DAN Addition of PAB I, but not deadenylation of the RNA and not PAB I is a but to the for the PAB M. G. Cell. 1994; PubMed Scopus Google Scholar, T. J. Mol. Biol. 1996; PubMed Scopus Google Scholar). In the of PAB I, strongly of decay intermediates differed in length by about 30 to those in cytoplasmic extract. The reaction about 15 A at the 3′-end. or deadenylated RNA was The pattern of deadenylation intermediates in the reconstituted is shown by with that in cytoplasmic extract in The PAB reaction was the At a ratio of DAN and deadenylation was within in the of spermidine In the of PAB I, deadenylation was not after of nuclease was used the poly(A)-binding protein (PAB A. P. P. J. Biol. Chem. 1993; Full Text PDF PubMed Google was used at the same concentration as PAB I, DAN was not were carried out at spermidine-independent deadenylation as the deadenylated RNA was Under these conditions, PAB I deadenylation in a At the concentration of PAB I used to PAB activity low salt was At PAB I deadenylation was and intermediates those However, the prominent deadenylation about A rather and deadenylated RNA As of DAN were stimulated by the purified enzyme was strongly The spermidine the on the the DAN were to spermidine-independent activity with purified were more in the of DAN PAB I at concentration and that only of this protein not Thus, a protein is responsible for this activity. by of two the of which the to and a as are a to for attack on the The of this reaction are two in a 3′-OH and a Biol. Scopus Google J. Biol. Chem. 1993; Full Text PDF PubMed Google Scholar). of and requirement for a 3′-OH at the DAN to the have been purified from eucaryotic A poly(A)-specific 3′-exoribonuclease has been and purified from HeLa cell extract J. A. A. J. PubMed Scopus Google Scholar, J. A. A. J. Biol. Chem. 1992; Full Text PDF PubMed Google Scholar). in properties and suggest that may have purified this nuclease. A Mg2+-dependent 3′-exoribonuclease has been purified from enzyme also and requires a free 3′-OH Ross J. J. Biol. Chem. 1994; Full Text PDF PubMed Google Scholar). it has a of kDa and is not it is not to In yeast, a 3′-exoribonuclease activity is with the of the essential involved in P. D. Genes Dev. 1996; PubMed Scopus Google Scholar). The activity requires a free and eucaryotic described have been by J. A. A. J. Biol. Chem. 1992; Full Text PDF PubMed Google Scholar, P. D. Genes Dev. 1996; PubMed Scopus Google Scholar). physiological DAN to be to the poly(A) PAN (11Sachs A.B. Deardorff J.A. Cell. 1992; 70: 961-973Abstract Full Text PDF PubMed Scopus (160) Google Scholar, 12Lowell J.E. Rudner D.Z. Sachs A.B. Genes Dev. 1992; 6: 2088-2099Crossref PubMed Scopus (91) Google Scholar). DAN and PAN are Mg2+-dependent poly(A)-specific that a free 3′-OH group, as and degrade poly(A) in a distributive The common to PAN and DAN, is the on RNA-binding like PAN, the activity was in the of PAB A PAN and PAB I has been R. Tarun Jr., S. Rieger M. Deardorff J.A. Müller-Auer S. Sachs A.B. J. Biol. Chem. 1996; 271: 432-438Abstract Full Text Full Text PDF PubMed Scopus (147) Google Scholar). DAN was stimulated by PAB I only and yet were more the of DAN on these is at low salt concentration, the physiological to be in poly(A)-specific from the that the first and rate-limiting step for the decay of many mRNAs in as as in higher eucaryotes is poly(A) shortening (8Jacobson A. Peltz S.W. Annu. Rev. Biochem. 1996; 65: 693-739Crossref PubMed Scopus (577) Google Scholar). The role of PAB I in this process is As in suggest a PAB I requirement for deadenylation. the of the enzyme PAN on PAB I is in agreement with the the properties of and mutants have not that the enzyme is involved in deadenylation of vivo R. Tarun Jr., S. Rieger M. Deardorff J.A. Müller-Auer S. Sachs A.B. J. Biol. Chem. 1996; 271: 432-438Abstract Full Text Full Text PDF PubMed Scopus (147) Google Scholar, 14Brown C.E. Tarun S.Z. Boeck R. Sachs A.B. Mol. Cell. Biol. 1996; 16: 5744-5753Crossref PubMed Scopus (130) Google Scholar). In has been that PAB I may be of deadenylation (18Bernstein P. Peltz S.W. Ross J. Mol. Cell. Biol. 1989; 9: 659-670Crossref PubMed Scopus (297) Google Scholar, M. J. 1996; 15: PubMed Scopus Google Scholar). The that DAN can be either or stimulated by PAB I, on the reaction conditions, that the the PAB I of deadenylation in and a PAB I of the same reaction in may not be as as might PAB I is or it is that the for the deadenylation reaction is not naked poly(A) in cells, a of poly(A) and PAB is suggested by the that PAB I is in to poly(A) in the M. G. Cell. 1994; PubMed Scopus Google Scholar). the pattern of deadenylation intermediates in vivo is by the properties of PAB I J. Cell. Biol. PubMed Scopus Google Scholar). have a deadenylation reaction in cytoplasmic extract that the same pattern of intermediates as in vivo. The reaction reconstituted with purified DAN and PAB I the same were which deadenylation was PAB the of the reaction about 15 A is to the poly(A) to PAB I A.B. Davis R.W. Mol. Cell. Biol. 7: PubMed Scopus Google Scholar). or deadenylated RNA was A of this pattern be that PAB I has a is required for to DAN, but the is protected At higher salt concentration, deadenylation was by PAB Under of a pattern of intermediates was However, the tail a length of about corresponding to the protected by PAB I J. Cell. Biol. PubMed Scopus Google and deadenylated RNA was the Under these conditions, to the RNA to DAN, the pattern to be to of poly(A) by PAB the effect is these may in or of either PAB I or may that the pattern of intermediates is a PAB I and be with However, in with the 3′-exonuclease pattern was DAN was the poly(A) degrading activity in cell The properties of the enzyme suggest that it plays role in poly(A) tail in vivo. experiments whether this role is in the deadenylation reaction that to mRNA for the and for with and for to the of the of for the and and for also and for with Sachs for and for in the are to for the time this was at the of the of of