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Journal of Bacteriology, July 2005, p. 4698-4706, Vol. 187, No. 14
0021-9193/05/$08.00+0     doi:10.1128/JB.187.14.4698-4706.2005
Copyright © 2005, American Society for Microbiology. All Rights Reserved.

Addition of Poly(A) and Heteropolymeric 3' Ends in Bacillus subtilis Wild-Type and Polynucleotide Phosphorylase-Deficient Strains

Departamento de Ingeniería Genética de Plantas, Centro de Investigación y de Estudios Avanzados, Unidad Irapuato, Irapuato, México,¹ Department of Biology, Emory University, Atlanta, Georgia 30322,² Department of Pharmacology and Biological Chemistry, Mount Sinai School of Medicine, New York University, New York, New York 10029³

Received 24 February 2005/ Accepted 19 April 2005

Polyadenylation plays a role in decay of some bacterial mRNAs, as well as in the quality control of stable RNA. In Escherichia coli, poly(A) polymerase I (PAP I) is the main polyadenylating enzyme, but the addition of 3' tails also occurs in the absence of PAP I via the synthetic activity of polynucleotide phosphorylase (PNPase). The nature of 3'-tail addition in Bacillus subtilis, which lacks an identifiable PAP I homologue, was studied. Sizing of poly(A) sequences revealed a similar pattern in wild-type and PNPase-deficient strains. Sequencing of 152 cloned cDNAs, representing 3'-end sequences of nontranslated and translated RNAs, revealed modified ends mostly on incomplete transcripts, which are likely to be decay intermediates. The 3'-end additions consisted of either short poly(A) sequences or longer heteropolymeric ends with a mean size of about 40 nucleotides. Interestingly, multiple independent clones exhibited complex heteropolymeric ends of very similar but not identical nucleotide sequences. Similar polyadenylated and heteropolymeric ends were observed at 3' ends of RNA isolated from wild-type and pnpA mutant strains. These data demonstrated that, unlike the case of some other bacterial species and chloroplasts, PNPase of Bacillus subtilis is not the major enzyme responsible for the addition of nucleotides to RNA 3' ends.

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