Barcoded reciprocal hemizygosity analysis via sequencing illuminates the complex genetic basis of yeast thermotolerance.

Decades of successes in statistical genetics have revealed the molecular underpinnings of traits as they vary across individuals of a given species. But standard methods in the field cannot be applied to divergences between reproductively isolated taxa. Genome-wide reciprocal hemizygosity mapping (RH-seq), a mutagenesis screen in an interspecies hybrid background, holds promise as a method to accelerate the progress of interspecies genetics research. Here, we describe an improvement to RH-seq in which mutants harbor barcodes for cheap and straightforward sequencing after selection in a condition of interest. As a proof of concept for the new tool, we carried out genetic dissection of the difference in thermotolerance between two reproductively isolated budding yeast species. Experimental screening identified dozens of candidate loci at which variation between the species contributed to the thermotolerance trait. Hits were enriched for mitosis genes and other housekeeping factors, and among them were multiple loci with robust sequence signatures of positive selection. Together, these results shed new light on the mechanisms by which evolution solved the problems of cell survival and division at high temperature in the yeast clade, and they illustrate the power of the barcoded RH-seq approach.

Joint effects of genes underlying a temperature specialization tradeoff in yeast.

A central goal of evolutionary genetics is to understand, at the molecular level, how organisms adapt to their environments. For a given trait, the answer often involves the acquisition of variants at unlinked sites across the genome. Genomic methods have achieved landmark successes in pinpointing these adaptive loci. To figure out how a suite of adaptive alleles work together, and to what extent they can reconstitute the phenotype of interest, requires their transfer into an exogenous background. We studied the joint effect of adaptive, gain-of-function thermotolerance alleles at eight unlinked genes from Saccharomyces cerevisiae, when introduced into a thermosensitive sister species, S. paradoxus. Although the loci damped each other's beneficial impact (that is, they were subject to negative epistasis), most boosted high-temperature growth alone and in combination, and none was deleterious. The complete set of eight genes was sufficient to confer ~15% of the S. cerevisiae thermotolerance phenotype in the S. paradoxus background. The same loci also contributed to a heretofore unknown advantage in cold growth by S. paradoxus. Together, our data establish temperature resistance in yeasts as a model case of a genetically complex evolutionary tradeoff, which can be partly reconstituted from the sequential assembly of unlinked underlying loci.

Population and comparative genetics of thermotolerance divergence between yeast species.

Many familiar traits in the natural world-from lions' manes to the longevity of bristlecone pine trees-arose in the distant past, and have long since fixed in their respective species. A key challenge in evolutionary genetics is to figure out how and why species-defining traits have come to be. We used the thermotolerance growth advantage of the yeast Saccharomyces cerevisiae over its sister species Saccharomyces paradoxus as a model for addressing these questions. Analyzing loci at which the S. cerevisiae allele promotes thermotolerance, we detected robust evidence for positive selection, including amino acid divergence between the species and conservation within S. cerevisiae populations. Because such signatures were particularly strong at the chromosome segregation gene ESP1, we used this locus as a case study for focused mechanistic follow-up. Experiments revealed that, in culture at high temperature, the S. paradoxus ESP1 allele conferred a qualitative defect in biomass accumulation and cell division relative to the S. cerevisiae allele. Only genetic divergence in the ESP1 coding region mattered phenotypically, with no functional impact detectable from the promoter. Our data support a model in which an ancient ancestor of S. cerevisiae, under selection to boost viability at high temperature, acquired amino acid variants at ESP1 and many other loci, which have been constrained since then. Complex adaptations of this type hold promise as a paradigm for interspecies genetics, especially in deeply diverged traits that may have taken millions of years to evolve.

Non-microvascular Successful Management of Near-total Ear Avulsion.

Traumatic ear avulsion represents a difficult challenge for all reconstructive surgeons; hence, replantation and reattachment will provide the best aesthetic outcome. However, when microsurgery is not possible, the surgeon must choose the most appropriate alternative method to address the concern. We are reporting a case of a near-total right ear external auricle amputation attached only by small skin bridge. The patient was admitted to our institute 22 hours after sustaining an injury during a motor vehicle collision. He was successfully managed by reattaching the external auricle into the anatomical place, with the repair of cartilage. This was followed by applying a daily protocol for venous congestion with the use of subcutaneous and intradermal injection of low-molecular-weight heparin daily for 10 days with gradual tapering of the dose. The external auricle survived with no complications or morbidity. Subcutaneous and intradermal low-molecular-weight heparin can be used effectively in cases of severe venous congestion of avulsed ear with adequate arterial inflow without causing any morbidity.