Davidson, M. K. | bioRxiv Science

bioRxiv2024

Laboratory horror stories: Poison in the agars

The fission yeast Schizosaccharomyces pombe is a single-celled eukaryote that can be cultured as a haploid or as a diploid. Scientists employ mating, meiosis, and the plating of ascospores and cells to generate strains with novel genotypes and to discover biological processes. Our two laboratories encountered independently sudden-onset, major impediments to such research. Spore suspensions and vegetative cells no longer plated effectively on minimal media. By systematically analyzing multiple different media components from multiple different suppliers, we identified the source of the problem. Specific lots of agar, from different suppliers, were toxic. Interestingly, the inhibitory effect was attenuated on rich media. Consequently, quality control checks that use only rich media can provide false assurances on the quality of the agar. Lastly, we describe likely sources of the toxicity and we provide specific guidance for quality control measures that should be applied by all vendors as preconditions for their sale of agar. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=116 SRC="FIGDIR/small/597796v1_ufig1.gif" ALT="Figure 1"> View larger version (40K): org.highwire.dtl.DTLVardef@41d7c1org.highwire.dtl.DTLVardef@bbec24org.highwire.dtl.DTLVardef@18c6f76org.highwire.dtl.DTLVardef@e9b707_HPS_FORMAT_FIGEXP M_FIG C_FIG Take-awayO_LISporadically, batches of agar from different suppliers strongly inhibit the plating efficiency of S. pombe spores and vegetative cells on minimal media. C_LIO_LIQuality control checks that are not quantitative or that use only rich media can provide false assurances on the quality of the agar. C_LIO_LIVendors should conduct rigorous, thorough, organism-specific tests for potential toxicity of each lot of agar as a pre-condition for its sale. C_LI

microbiology↗

bioRxiv2020

Molecular mechanisms for environmentally induced plasticity in the positioning of meiotic recombination at hotspots

In meiosis, Spo11/Rec12-initiated homologous recombination is clustered at hotspots that regulate its frequency and distribution across the genome. Intriguingly, the intensities and positions of recombination hotspots can change dramatically in response to intracellular and extracellular conditions, and can display epigenetic memory. Here, using the fission yeast Schizosaccharomyces pombe, we reveal mechanisms for hotspot plasticity. We show that each of six hotspot-activating proteins (transcription factors Atf1, Pcr1, Php2, Php3, Php5, Rst2) is rate-limiting for promoting recombination at its own DNA binding site, allowing each class of hotspot to be regulated independently by agonistic and antagonistic signals. We also discovered that the regulatory protein-DNA complexes can establish a recombinationally poised epigenetic state before meiosis. Notably, Atf1 and Pcr1 controlled the activation of DNA sequence-dependent hotspots to which they do not bind; and they do so by regulating the expression of other hotspot-activating proteins. Thus, while each transcription factor activates its own class of DNA sequence-dependent hotspots directly in cis, cross-talk between regulatory networks modulates in trans the frequency and positioning of recombination at other classes of DNA sequence-dependent hotspots. We posit that such mechanisms allow cells to alter the frequency distribution of meiotic recombination in response to metabolic states and environmental cues.

genetics↗