Volume 19 Part 1 Article 70
Title: Interruption of a MSH4 Homolog in Pleurotus ostreatus is Responsible for the Absence of Spore Production
Author: Brian Lavrijssen, Johan JP Baars, Anton SM Sonnenberg
The sporeless oyster mushroom (Pleurotus ostreatus) variety SPOPPO, introduced to the market in 2006, has been generated by using an existing sporeless mutant (ATCC58937) as the donor and the commercial variety HK35 as acceptor strain. The sporeless phenotype appeared to be recessive and was mapped in both constituent nuclei on the same locus, suggesting the involvement of one gene. The present research was directed to identify the gene(s) involved, thus facilitating breeding additional sporeless varieties and see if this gene can be a target for breeding sporeless varieties in other edible fungi. The locus containing the sporeless trait was fine mapped to 78.9 kb containing 27 predicted genes. The equivalent region of the sequenced sporulating strain N001 PC15 was subcloned as individual genes or pairs of genes and used to transform the non-sporulating mutant. One gene was able to restore the sporulation completely. The commercial sporeless variety shows, next to the absence of spores, also the loss of orientation of fruiting body in each bunch indicating a disturbance of negative gravitropism. With restoration of sporulation, also the negative gravitropism of the fruiting bodies was restored in the transformants, indicating the involvement of the gene also in this phenotype. The 3,864 bp gene encoding an 852 amino acid protein was identified as a MutS protein homolog 4 (MSH4) homolog. MSH4 is known to be meiosis specific and required for reciprocal recombination and proper segregation of homologous chromosomes at meiosis I. The cDNA sequence of the wild type of MSH4 revealed 26 introns ranging in size between 8 and 300 bp. Comparing the coding regions of the wild type and the mutant MSH4 homologs, the latter constructed from the Illumina reads, showed two amino acid shifts in the mutant that do not seem to be located in positions crucial for its function. A de novo assembly of one of the homokaryons of the non-sporulating strain indicates that the gene is interrupted, possibly by a 5 kb sequence. Due to the poor quality of the de novo assembly, only part of the sequence of insertion is known. A BLAST search on the whole genome indicates that it represents a repetitive sequence, possibly a LTR-transposon.