Beaudry, R. | Search bioRxiv

bioRxiv2023

Apple and banana fruits produce anteiso- and iso-branched-chain esters from newly synthesized precursors

Inhibitors of acetohydroxyacid synthase, the common enzyme of branched-chain amino acid biosynthesis, were applied to ripening apple (Malus xdomestica Borkh.), banana (Musa spp.), and flowering quince (Chaenomeles xsuperba) fruits to discern the contribution of newly synthesized precursors to branched-chain ester formation. After treatment, anteiso- and iso-branched-chain volatiles (i.e., those related to isoleucine, and valine and leucine, respectively) were observed to universally decrease in content. Fruits recovered production following exogenous feeding of branched-chain -ketoacids. Furthermore, apple and banana fruits were capable of metabolizing all three branched-chain -ketoacids to esters. Among free amino acids, only the branched-chain amino acids with correspondingly reduced branched-chain esters had a lesser concentration following treatment with inhibitor. Our results ultimately reject the hypothesis that anteiso- and iso-branched-chain esters are derived from preexisting amino acids and instead support the hypothesis that these esters are the product of de novo precursor biosynthesis. The novel use of these inhibitors also allowed for further investigation of branched-chain volatile biosynthesis, the citramalate synthase pathway, and the importance of precursor availability in fruits. Notably, in Valery banana fruit, ethyl acetate and butyl acetate were found to be dependent on acetohydroxyacid synthase activity for production whereas 1-methylbutyl acetate and 1-methylbutyl butanoate (sec-branched-chain esters) were not. Inhibitor usage on apples also allowed for a sensory study that found that humans can discern the absence of 2-methylbutyl and 2-methylbutanoate esters in apple fruit. Additionally, a population genetics analysis found that there is selection pressure against apples that lack these esters.

biochemistry↗

bioRxiv2023

The identification and analysis of meristematic mutations within the apple tree that developed the RubyMac sport mutation

Understanding the molecular basis of sport mutations in fruit trees can accelerate breeding of novel cultivars. For this, we analyzed the DNA of the apple tree that evolved the RubyMac phenotype through a sport mutation that introduced changes in fruit coloration in upper branches of the tree. Unexpectedly, we not only found 46 de novo mutations, but also 54 somatic gene conversions (i.e., loss-of-heterozygosity mutations) distinguishing the mutant and wild-type branches of the tree. Approximately 30% of the de novo mutations and 80% of the gene conversions were observed only in specific cells layers suggesting that they occurred in the corresponding meristematic layers. Interestingly, the de novo mutations were enriched for GC=>AT transitions, while the gene conversions showed the opposite bias for AT=>GC transitions suggesting that GC-biased gene conversions have the potential to counteract the AT-bias of de novo mutations. By comparing the gene expression patterns in fruit skins from mutant and wild-type branches, we found 56 differentially expressed genes including 18 that were involved in anthocyanin biosynthesis. While none of the differently expressed genes harbored a mutation, we found that some of the mutations affected the integrity of candidate genes in regions of the genome that were recently associated with natural variation in fruit coloration.

genomics↗