Polynucleotide phosphorylase Pnp and exonuclease Rnr, isolated by specific sorption on nucleic acids, process RNA products derived from «promoter islands»

Abstract

Oligonucletides synthesized from promoter-rich regions of the bacterial genome («promoter islands
») were studied. Despite multiple sites for potential transcription initiation in their sequences, short oligonucleotides
rather than full-size RNAs are the main products originating from the “islands”. This anomaly
may be due to the peculiarities of the structural and functional organization of the «islands» and/or targeted
post-transcriptional processing of synthesized products. To assess the second possibility, a fraction of purified
biotynilated short RNAs of Escherichia coli was immobilized on magnetic beads and used for specific
sorption of proteins from bacterial cell lysate. LC/MS spectrometry revealed a number of proteins capable of
strong binding to small RNAs, and among them was polynucleotide phosphorilase Pnp. The potentiality of
this nuclease and RNase Rnr, which was identified previously among proteins specifically sorbed on fragments
of «promoter islands», to affect the intracellular contents of short and long «island»-derived RNAs was
studied by real-time RT-PCR. It turned out that rnr deletion results in an increase in the amount of all studied
types of RNA in bacterial cells, while deletion of pnp affected the intracellular level of three out of four model
samples. Thus, posttranscriptional processing plays an important role in the genesis of “island” RNA products,
which can realize certain functions, yet to be elucidated.