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.2011 Feb;193(3):734-43.
doi: 10.1128/JB.01016-10. Epub 2010 Nov 19.

Genome copy numbers and gene conversion in methanogenic archaea

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Genome copy numbers and gene conversion in methanogenic archaea

Catherina Hildenbrand et al. J Bacteriol.2011 Feb.

Abstract

Previous studies revealed that one species of methanogenic archaea, Methanocaldococcus jannaschii, is polyploid, while a second species, Methanothermobacter thermoautotrophicus, is diploid. To further investigate the distribution of ploidy in methanogenic archaea, species of two additional genera-Methanosarcina acetivorans and Methanococcus maripaludis-were investigated. M. acetivorans was found to be polyploid during fast growth (t(D) = 6 h; 17 genome copies) and oligoploid during slow growth (doubling time = 49 h; 3 genome copies). M. maripaludis has the highest ploidy level found for any archaeal species, with up to 55 genome copies in exponential phase and ca. 30 in stationary phase. A compilation of archaeal species with quantified ploidy levels reveals a clear dichotomy between Euryarchaeota and Crenarchaeota: none of seven euryarchaeal species of six genera is monoploid (haploid), while, in contrast, all six crenarchaeal species of four genera are monoploid, indicating significant genetic differences between these two kingdoms. Polyploidy in asexual species should lead to accumulation of inactivating mutations until the number of intact chromosomes per cell drops to zero (called "Muller's ratchet"). A mechanism to equalize the genome copies, such as gene conversion, would counteract this phenomenon. Making use of a previously constructed heterozygous mutant strain of the polyploid M. maripaludis we could show that in the absence of selection very fast equalization of genomes in M. maripaludis took place probably via a gene conversion mechanism. In addition, it was shown that the velocity of this phenomenon is inversely correlated to the strength of selection.

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Figures

FIG. 1.
FIG. 1.
Overview of the method for the quantification of genome copy numbers. Details are explained in the text.
FIG. 2.
FIG. 2.
Growth-phase-dependent ploidy of fast-growingM. acetivorans. (A) An average growth curve ofM. acetivorans grown on methanol is shown that was derived from five independent cultures. Standard deviations are included. Numbers and arrows indicate the times at which aliquots were removed for quantification of the genome copy numbers. The arithmetic instead of the semilogarithmic representation (which is standard for physiological experiments concentrating on exponential phase) was chosen to better visualize the transition between exponential and stationary phase. (B) Genome copies per cell ofM. acetivorans grown on methanol. Average values and SDs from five independent experiments for the following growth phases are shown: 1, early exponential phase; 2, mid-exponential phase; 3, late exponential phase; 4, early stationary phase; and 5, late stationary phase.
FIG. 3.
FIG. 3.
Growth-phase-dependent ploidy of slow-growingM. acetivorans. (A) An average growth curve ofM. acetivorans grown on acetate is shown that was derived from five independent cultures. For further explanation, see the legend to Fig. 2. (B) Genome copies per cell ofM. acetivorans grown on sodium acetate. For further explanation, see the legend to Fig. 2.
FIG. 4.
FIG. 4.
Growth-phase-dependent ploidy ofM. maripaludis. (A) An average growth curve ofM. maripaludis grown on formate is shown that was derived from five independent cultures. For further explanation, see the legend to Fig. 2. (B) Genome copies per cell ofM. maripaludis grown on sodium formate. Average values and SDs from five independent experiments for the following growth phases are shown: 1, early exponential phase; 2, late exponential phase; 3, early stationary phase; and 4, late stationary phase.
FIG. 5.
FIG. 5.
Gene conversion under different selection pressures. A strain simultaneously containing genomes with theselD and thepacN gene at theselD locus was precultured under full selection pressure (2.5 μg of puromycin/ml) to minimize the number ofselD-containing genomes. The preculture was used to inoculate four cultures, which were incubated under three different levels of selection (+++, 2.5 μg of puromycin/ml; ++, 0.6 μg of puromycin/ml; and +, 0.15 μg of puromycin/ml) and in the absence of selection (−). The four cultures were grown for ∼20 generations via consecutive dilution into fresh medium. After the first-, eighth-, and fourteenth-generation aliquots were withdrawn, the copy numbers of both types of genomes were determined by real-time PCR. Average values and SDs are shown. M,pacN-containing mutant genome; WT,selD-containing wild-type genome.
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References

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