VIEW Open Access Two Genomes Are Better Than One: History, Genetics, and Biotechnological Applications of Fungal Heterokaryons Noah B
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Strom and Bushley Fungal Biol Biotechnol (2016) 3:4 DOI 10.1186/s40694-016-0022-x Fungal Biology and Biotechnology REVIEW Open Access Two genomes are better than one: history, genetics, and biotechnological applications of fungal heterokaryons Noah B. Strom* and Kathryn E. Bushley Abstract Heterokaryosis is an integral part of the parasexual cycle used by predominantly asexual fungi to introduce and main- tain genetic variation in populations. Research into fungal heterokaryons began in 1912 and continues to the present day. Heterokaryosis may play a role in the ability of fungi to respond to their environment, including the adaptation of arbuscular mycorrhizal fungi to different plant hosts. The parasexual cycle has enabled advances in fungal genetics, including gene mapping and tests of complementation, dominance, and vegetative compatibility in predominantly asexual fungi. Knowledge of vegetative compatibility groups has facilitated population genetic studies and enabled the design of innovative methods of biocontrol. The vegetative incompatibility response has the potential to be used as a model system to study biological aspects of some human diseases, including neurodegenerative diseases and cancer. By combining distinct traits through the formation of artificial heterokaryons, fungal strains with superior properties for antibiotic and enzyme production, fermentation, biocontrol, and bioremediation have been produced. Future biotechnological applications may include site-specific biocontrol or bioremediation and the production of novel pharmaceuticals. Keywords: Heterokaryon, Parasexual cycle, Complementation, Vegetative compatibility groups, Heterosis, Protoplast fusion, Biotechnology Background of ascogenous cells [2]. In the sexual cycle, the nuclei in Heterokaryosis refers to the presence of two or more these dikaryotic cells fuse and undergo meiosis, result- genetically distinct nuclei within the same cell. While ing in genetically recombined haploid basidiospores or uncommon throughout most of life’s kingdoms, het- ascospores. erokaryosis is a hallmark of kingdom Fungi. The fungal Heterokaryosis also occurs during vegetative growth subkingdom, Dikarya, which contains two phyla (Asco- as a component of the parasexual cycle, a mechanism for mycota and Basidiomycota) and 95 % of all known fungal increasing genotypic diversity in predominantly asexual species [1], is named for its characteristic heterokaryons fungi (Fig. 1) [3]. In this paper, the term “predominantly with exactly two genetically distinct nuclei. Basidiomy- asexual” is used to refer to fungi that historically were not cota are often distinguished by the presence of clamp known to have a sexual cycle and those that only rarely connections, a cytological feature unique to this phy- undergo sexual recombination. In the parasexual cycle, lum that helps to ensure the segregation of both haploid first described in 1956 by Pontecorvo [4], hyphae from nuclei into daughter cells following mitosis. Ascomycetes two compatible individuals grow towards each other by are often characterized by croziers, structures similar to chemotaxis [5] and fuse, allowing exchange of nuclei to clamp connections, that maintain the dikaryotic state form a heterokaryon [4]. These nuclei become mixed in the cytoplasm of the heterokaryon [6] and sometimes undergo karyogamy, resulting in diploid cells. Following *Correspondence: [email protected] Department of Plant Biology, University of Minnesota, 826 Biological karyogamy, mitotic recombination and repeated chro- Sciences, 1445 Gortner Avenue, Saint Paul, MN 55108, USA mosome loss through mitotic nondisjunction result in © 2016 Strom and Bushley. This article is distributed under the terms of the Creative Commons Attribution 4.0 International License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution, and reproduction in any medium, provided you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The Creative Commons Public Domain Dedication waiver (http://creativecommons. org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated. Strom and Bushley Fungal Biol Biotechnol (2016) 3:4 Page 2 of 14 a create beneficial fungal hybrids in applications as diverse as antibiotic production and bioremediation (Fig. 2). This review focuses on the evolution of scientific thought about non-sexual heterokaryosis, its biotechnological b applications, and future directions for basic and applied research. Early discoveries c The term, heterokaryosis, was coined in 1912 by Ger- man mycologist, Hans Burgeff [8]. Working with Phyco- myces nitens, he noticed that different sporangiospores d could give rise to phenotypically distinct mycelia [8, 9], contrary to the phenotypically identical offspring one would predict from asexual mitotic reproduction. In 1932, Hansen and Smith, working with Botrytis cinerea, showed that the phenotypically distinct mycelia arose from either one or the other haploid nucleus or from both, which were variably present in the sporangiospores e [10]. Their explanation, now known to be correct, was that dissimilar nuclei of the heterokaryotic mycelium segregated into different cells, resulting in homokaryotic regions of the mycelium that differed from each other in their genetic makeup. In 1944, Beadle and Coonradt, working with Neuro- spora crassa, produced two auxotrophic strains, one of which could not synthesize p-aminobenzoic acid, and the other of which could not synthesize nicotinic acid [11]. When grown separately on minimal media, neither strain Fig. 1 The parasexual cycle. The parasexual cycle parallels events in the sexual cycle, resulting in genetically unique haploid offspring thrived, but when grown together on minimal media, but without a meiotic reduction. a Hyphae of genetically unique hyphal fusion allowed for the formation and growth of a homokaryotic parents grow towards each other by chemotaxis and heterokaryon possessing a functional copy of each gene fuse. b Nuclei from each unique strain migrate within the fused deficient in the other strain. This genetic complemen- hypha, which is now considered a heterokaryon. c Haploid nuclei tation experiment suggested the potential advantages in the heterokaryon undergo karyogamy to create a heterozygous diploid nucleus. d The diploid nucleus undergoes mitotic recombi- of heterokaryosis in wild asexual fungi and a rationale nation to produce a recombined genotype. e In growing hyphae, for the persistence of heterokaryons throughout fungal gradual loss of chromosomes due to repeated rounds of mitotic non- evolution. disjunction results in haploidization and unique genotypes in various sectors of mycelium Heterokaryons and adaptation Beadle and Coonradt also suggested that nuclear exchanges may take place multiple times in the lifetime of a single fungus, allowing fungi to continually update the formation of haploid, as well as some aneuploid, cells their genomes through contact with compatible neigh- with unique genomes from those of either parent nucleus bors [11]. This potential for genetic variation within the [4]. Thus, even in the absence of meiosis and sexual lifetime of a single fungus was hypothesized to play an reproduction, the parasexual cycle is effective in increas- important role in the fungus’ ability to adapt to a con- ing genotypic diversity in predominantly asexual fungi tinuously changing environment. In 1952, Jinks provided [7]. the first experimental data to support the hypothesis that Fungal biologists have taken advantage of heterokaryo- genotypically distinct nuclei within the same mycelium sis during the parasexual cycle to enable genetic analyses could be involved in adaption [12]. In a culturing experi- and to advance biotechnology. For example, heterokary- ment involving various types of media, he showed that ons have enabled gene mapping, complementation, and the ratios of two genotypically unique nuclei within a wild epidemiological studies in predominantly asexual fungi. strain of Penicillium fluctuated according to media type. The power of heterokaryosis has also been harnessed to Similar unbalanced ratios of nuclei have been observed Strom and Bushley Fungal Biol Biotechnol (2016) 3:4 Page 3 of 14 a Cell wall digestion with cell wall degrading enzymes b PEG-induced protoplast fusion c Protoplast regeneration d e Genetic analyses Fermentation Pharmaceutical productionBioremediation Biocontrol Fig. 2 Protoplast fusion. a Fungal cell walls are digested with cell wall degrading enzymes, exposing protoplasts (b). c Polyethylene glycol (PEG) facilitates fusion of protoplasts, which may belong to different strains or species, as represented by red or blue chromosomes. d Fused protoplasts form heterokaryons and may progress partway or completely through the parasexual cycle, resulting in diploid or recombined haploid hybrids. e Applications of fungal heterokaryons or hybrids resulting from protoplast fusion include genetic analyses, fermentation, pharmaceutical production, bioremediation, and biocontrol in the basidiomycete, Heterobasidion parviporum [13]. remains controversial as to whether AMF, ubiquitous Whether these shifts resulted from selection acting on plant root symbionts whose spores are multinucleate nuclei [14] or from more random processes of gain or [22], are homokaryotic or heterokaryons formed through loss in particular mycelial fractions