Genetic basis of the complementary Dpnl and Dpnll restriction systems of S. pneumoniae: An intercellular cassette mechanism

doi:10.1016/0092-8674(86)90698-7

Cell

Volume 46, Issue 7, 26 September 1986, Pages 993-1000

https://doi.org/10.1016/0092-8674(86)90698-7 Get rights and content

Abstract

Cells of S. pneumoniae contain either Dpnl, a restriction endonuclease that cleaves only the methylated DNA sequence 5′-GmeATC-3′, or Dpnll, which cleaves the same sequence when not methylated. A chromosomal DNA segment containing Dpnll genes was cloned in S. pneumoniae. Nucleotide sequencing of this segment revealed genes encoding the methylase and endonuclease and a third protein of unknown function. When the plasmid was introduced into Dpnl cells, recombination during chromosomal facilitation of its establishment substituted genes encoding the Dpnl endonuclease and another protein in place of the Dpnll genes. DNA hybridization and sequencing showed that the Dpnl and Dpnll segments share homology on either side but not between themselves or with other regions of the chromosome. Thus, the complementary restriction systems are found on non-homologous and mutually exclusive cassettes that can be inserted into a particular point in the chromosome of S. pneumoniae on the basis of neighboring homology.

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Cited by (92)

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Citation Excerpt :
Among them, the type Ⅰ-Ⅲ system is consisted of methyltransferase (MTase) which methylates the self-DNA and restrictase (REase) which degrades foreign DNA [15,16], while the type Ⅳ R-M system contains only a DNA modification-dependent REase which degrades the corresponding DNA with epigenetic modification [14]. For example, the type Ⅳ REase Dpn Ⅰ, which has been widely used in site-directed mutagenesis [17], degrades foreign DNA with 5′-GmATC-3' (adenine is methylated) in Streptococcus pneumonia [18–21]. Some bacteria and phages have only MTase which is called orphan MTase [22,23].
Accompanied by benefits from horizontally transferred genes, bacteria have to face the risk of the invasion of dangerous genes. Bacteria often use the restriction-modification (R-M) system, which is consisted of methyl transferase (MEase) and restrictase (REase), to protect self-DNA and defend against foreign DNA. Paenibacillus polymyxa, widely used as growth promoting rhizobacteria in agriculture, can also produce compounds of medical and industrial interests. It is unclear whether R-M systems exist in P. polymyxa. In this study, we used a shuttle plasmid with epigenetic modification from different bacteria to explore R-M systems in P. polymyxa. We found that DNA which is methylated by DNA adenine methyltransferase (Dam) in E. coli was strongly restricted, indicating the presence of a Dam-methylation-dependent R-M system in P. polymyxa. Whereas, DNA from a dam^- E. coli strain was also moderately restricted, indicating the presence of a Dam-methylation-independent R-M system. Degradation of plasmid DNA with Dam methylation by cell-free protein extract of P. polymyxa provides additional evidence for the presence of Dam-methylation-dependent R-M system. Taken together, our work showed that there are two different types of R-M system in P. polymyxa, providing a foundation for the study of innate immunity in P. polymyxa and for the development of genetic engineering tools in P. polymyxa.
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Streptomyces griseus does not readily take up foreign DNA isolated from other Streptomyces species or Escherichia coli, presumably due to its unique restriction-modification systems that function as a barrier for interspecific DNA transfer. To efficiently transform S. griseus by avoiding the restriction barriers, we methylated incoming DNA in vivo and in vitro and treated protoplasts with heat prior to transformation. Whereas heat treatment of protoplasts or methylation of the E. coli-Streptomyces shuttle vectors (pXE4 and pKK1443) did not prominently improve the transformation efficiency, HpaII methylation of the vectors from any E. coli strains tested in this study highly increased the transformation efficiency. The highest transformation efficiency was observed when the shuttle vectors were isolated from the dam, hsd strain of E. coli (GM161) and methylated by AluI and HpaII methyltransferases, and the efficiency was approximately the same as that of the vectors from S. griseus. We identified several restriction-modification systems that decrease the transformation efficiency. This research also led us to understand methylation profiles and restriction-modification systems in S. griseus.
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Cell

ArticleGenetic basis of the complementary Dpnl and Dpnll restriction systems of S. pneumoniae: An intercellular cassette mechanism

Abstract

Gene

Anal. Biochem.

Meth. Enzymol.

J. Biol. Chem.

J. Mol. Biol.

Meth. Enzymol.

J. Mol. Biol.

J. Mol. Biol.

Simultaneous production of two capsular polysaccharides by pneumococcus. II. The genetic and biochemical bases of binary capsulation

J. Exp. Med.

Heteroduplex DNA mismatch repair system of Streptococcus pneumoniae: cloning and expression of the hexA gene

J. Bacteriol.

Isolation of high molecular weight DNA from Hemophilus influenzae

J. Mol. Biol.

A rapid alkaline extraction procedure for screening recombinant DNA

Nucl. Acids Res.

Genetic control of restriction and modification in Escherichia coli

J. Bacteriol.

Identification of tetracycline resistant R-plasmids in Streptococcus agalactiae (group B)

Antimicrob. Ag. Chemother.

Interspecific plasmid transfer between Streptococcus pneumoniae and Bacillus subtilis

Mol. Gen. Genet.

Article
Genetic basis of the complementary Dpnl and Dpnll restriction systems of S. pneumoniae: An intercellular cassette mechanism