Trends in Microbiology
Volume 15, Issue 4, April 2007, Pages 172-180
Journal home page for Trends in Microbiology

Review
I will survive: DNA protection in bacterial spores

https://doi.org/10.1016/j.tim.2007.02.004Get rights and content

Dormant spores of Bacillus, Clostridium and related species can survive for years, largely because spore DNA is well protected against damage by many different agents. This DNA protection is partly a result of the high level of Ca2+-dipicolinic acid in spores and DNA repair during spore outgrowth, but is primarily caused by the saturation of spore DNA with a group of small, acid-soluble spore proteins (SASP), which are synthesized in the developing spore and then degraded after completion of spore germination. The structure of both DNA and SASP alters upon their association and this causes major changes in the chemical and photochemical reactivity of DNA.

Section snippets

Spore DNA resistance

Dormant spores of Bacillus and Clostridium species and their close relatives are formed in sporulation (Box 1; Box 2; Box 3) and can survive for hundreds of years and perhaps longer 1, 2, 3. The physiological properties of these spores are different from those of other known cells – Bacillus and Clostridium spores in water exhibit no detectable metabolism of endogenous or exogenous compounds and can survive treatments with wet and dry heat, UV and γ-radiation, desiccation and toxic chemicals

DNA protection in spores

The spore DNA is located in the core (Box 2), a region that has a lower water content (25–50% wet weight) than the protoplast of a growing cell or the outer spore layers (∼80% of wet weight is water in both) [13]. The spore core also has a pH of ∼6.5, which is ∼1 pH unit lower than in growing cells, and has a huge amount of pyridine-2,6-dicarboxylic acid [dipicolinic acid (DPA); ∼20% of core dry weight] 13, 14 (Box 1, Box 2). DPA in spores is almost certainly present as a 1:1 chelate with

α/β-type SASP

The α/β-type SASP are encoded by monocistronic genes, termed ssp, which are members of a multigene family of 3–7 members in spore-formers (Figure 1; Figure 2). The ssp genes are scattered on chromosomes (and on a plasmid in Bacillus cereus ATCC 10987; Figure 1), their promoters are upstream of and close to a strong ribosome-binding site, and coding sequences are followed by potential stem-loop structures that are probably transcription stop signals. Proteins encoded by ssp genes are not

Effects of α/β-type SASP on DNA properties in spores

Studies in vivo and in vitro have shown that α/β-type SASP have profound effects on DNA properties. Studies in vivo have largely used wild-type and αβB. subtilis spores. The αβ spores are significantly more sensitive than wild-type spores to wet and dry heat, UV radiation, dessication and genotoxic chemicals including nitrous acid, hydrogen peroxide and formaldehyde, although αβ and wild-type spores exhibit similar resistance to γ-radiation and DNA alkylating agents 4, 5, 9, 10, 24 (Table

Effects of α/β-type SASP on DNA in vitro and vice versa

Most of the effects of α/β-type SASP on DNA in spores are also seen in vitro using purified DNA and proteins, including α/β-type SASP from Bacillus and Clostridium species and proteins that are normally abundant and less abundant 4, 6, 7. All wild-type α/β-type SASP have similar effects on DNA in vitro, including slowing DNA depurination due to wet or dry heat, protecting DNA against cleavage by enzymes, hydroxyl radicals and hydrogen peroxide, preventing cytosine deamination to uracil and

Features of α/β-type SASP-DNA binding and the structure of the complex

All α/β-type SASP bind better to GC-rich DNAs, in particular to polydG–polydC, with polydA–polydT bound poorly if at all 14, 15, 34, 35, 39. dG6–dC6 is bound but binding is generally stronger to larger DNAs, especially for oligodATs. Mixed sequence DNAs such as plasmids can be saturated with α/β-type SASP, with apparent affinity constants as high as 107 M−1, although this value varies between different proteins 34, 35, 39. The site size for α/β-type SASP on DNA is 4–5 bp, binding is similar

Interaction of other components with α/β-type SASP-bound DNA in spores

Although the α/β-type SASP alone have striking effects on DNA properties in vitro, the situation in spores is likely to be more complex because additional components interact with DNA. These include the essential HBsu protein found at high levels on DNA in growing cells and spores [44], and Ca-DPA. HBsu decreases the persistence length of DNA saturated with α/β-type SASP in vitro without altering effects of α/β-type SASP on DNA photochemistry [44]. Perhaps the effect of HBsu on DNA persistence

Degradation of α/β-type SASP in spore outgrowth

Although α/β-type SASP are essential to spore DNA resistance, these proteins must be removed to enable DNA transcription in spore outgrowth. Two factors are important in protein removal: (i) dissociation of α/β-type SASP from DNA in fully germinated spores; and (ii) degradation of the proteins initiated by an endoprotease, Gpr, which is specific for α/β-type (and γ-type) SASP [45]. Peptidases then degrade the Gpr cleavage products to amino acids that support much protein synthesis and energy

Concluding remarks and future questions

The α/β-type SASP are the major factor protecting spore DNA from many damaging agents. These proteins are unique and, given their role in protecting DNA in a dormant organism where an entire chromosome is transcriptionally silent, this uniqueness might not be surprising. Indeed, the presence of these proteins in growing cells is extremely deleterious. Although much has been learned about these proteins and their interaction with DNA, several questions remain. These include the following: (i)

Acknowledgements

I thank all past and current members of the laboratory who have contributed to our understanding of the many aspects and effects of α/β-type SASP, Ca-DPA and Gpr, and to collaborators at other institutions, in particular Mark J. Jedrzejas and his coworkers. Work in my laboratory on these topics has been supported by grants from the National Institutes of Health (GM 19698) and the Army Research Office.

References (58)

  • R.J. Cano et al.

    Revival and identification of bacterial spores in 25- to 40-million-year-old Dominican amber

    Science

    (1995)
  • R.H. Vreeland

    Isolation of a 250 million-year old halotolerant bacterium from a primary salt crystal

    Nature

    (2000)
  • W.L. Nicholson

    Resistance of Bacillus endospores to extreme terrestrial and extraterrestrial environments

    Microbiol. Mol. Biol. Rev.

    (2000)
  • P. Setlow

    Spores of Bacillus subtilis: their resistance to radiation, heat and chemicals

    J. Appl. Microbiol.

    (2006)
  • P. Setlow

    Resistance of spores of Bacillus species to ultraviolet light

    Environ. Mol. Mutagen.

    (2001)
  • B. Setlow et al.

    Role of DNA repair in Bacillus subtilis spore resistance

    J. Bacteriol.

    (1996)
  • C.A. Loshon

    Formaldehyde kills spores of Bacillus subtilis by DNA damage, and small, acid-soluble spore proteins of the α/β-type protect spores against this DNA damage

    J. Appl. Microbiol.

    (1999)
  • R. Tennen

    Mechanisms of killing of spores of Bacillus subtilis by iodine, glutaraldehyde and nitrous acid

    J. Appl. Microbiol.

    (2000)
  • J.M. Salas-Pacheco

    Role of the Nfo (YqfS) and ExoA apurinic/apyrimidinic endonucleases in protecting Bacillus subtilis spores from DNA damage

    J. Bacteriol.

    (2005)
  • P. Gerhardt et al.

    Spore thermoresistance mechanisms

  • P. Setlow

    Mechanisms which contribute to the long-term survival of spores of Bacillus species.

    J. Appl. Bacteriol.

    (1994)
  • P. Setlow

    Mechanisms for the prevention of damage to the DNA in spores of Bacillus species

    Annu. Rev. Microbiol.

    (1995)
  • A. Driks

    Proteins of the spore core and coat

  • B. Setlow

    Role of dipicolinic acid in the resistance and stability of spores of Bacillus subtilis

    J. Bacteriol.

    (2006)
  • T. Douki

    Photosensitization of DNA by dipicolinic acid, a major component of spores of Bacillus species

    Photochem. Photobiol. Sci.

    (2005)
  • D. Raju

    Investigating the role of small, acid-soluble spore proteins (SASPs) in the resistance of Clostridium perfringens spores to heat

    BMC Microbiol.

    (2006)
  • Raju, D. et al. Antisense RNA-mediated decreased synthesis of small, acid-soluble spore proteins leads to decreased...
  • Y. Carrillo-Martinez et al.

    Properties of Bacillus subtilis small, acid-soluble, spore proteins with changes in the sequence recognized by their specific protease

    J. Bacteriol.

    (1994)
  • B. Setlow

    Effects of major spore-specific DNA binding proteins on Bacillus subtilis sporulation and spore properties

    J. Bacteriol.

    (2000)
  • Cited by (352)

    View all citing articles on Scopus
    View full text