Abstract
BackgroundThe ability to distinguish nonself from self is a fundamental characteristic of biological systems. In the filamentous fungus Neurospora crassa, multiple incompatibility genes mediate nonself recognition during vegetative growth. One of these genes, un-24, encodes both nonself recognition function and the large subunit of a type I ribonucleotide reductase, an evolutionarily conserved enzyme that is essential for the conversion of NDP precursors into dNDPs for use in DNA synthesis. Previous studies have shown that co-expression of the two allelic forms of un-24, Oakridge (OR) and Panama (PA), in the same cell results in cell death.ResultsWe identify a 135 amino acid nonself recognition domain in the C-terminus region of UN-24 that confers an incompatibility-like phenotype when expressed in the yeast, Saccharomyces cerevisiae. Low-level expression of this domain results in several cytological and phenotypic characteristics consistent with an incompatibility reaction in filamentous fungi. These incompatibility phenotypes are correlated with the presence of a non-reducible complex consisting of the PA incompatibility domain and Rnr1p, a large subunit of ribonucleotide reductase in yeast. When the PA incompatibility domain is switched to high-level expression, the incompatibility phenotype transitions to wild-type concomitant with the appearance of a complex containing the PA incompatibility domain and Ssa1p, an Hsp70 homolog.ConclusionsResults from this study provide insights into the mechanism and control of vegetative nonself recognition mediated by ribonucleotide reductase in N. crassa, thus establishing the yeast system as a powerful tool to study fungal nonself recognition. Our work shows that heat shock proteins may function to deactivate vegetative incompatibility systems, as required for entry into the sexual cycle. Finally, our results suggest that variations on the PA incompatibility domain may serve as novel and specific antimicrobial peptides.
Highlights
The ability to distinguish nonself from self is a fundamental characteristic of biological systems
Class I ribonucleotide reductase (RNR) are highly conserved across eukaryotes and operate as tetramers composed of two large subunits and two small subunits that catalyze the reduction of ribonucleoside diphosphates (NDPs) into deoxyribonucleoside diphosphates
Omission of six amino acids from the C-terminus [hygunPA(861– 917)] resulted in loss of incompatibility activity. Both specificity and incompatibility activity of UN-24PA is encompassed in a 135 amino acid domain that corresponds to the flexible C-terminus arm of the large subunit contained within the RNR large subunit found in yeast [13,14]
Summary
The ability to distinguish nonself from self is a fundamental characteristic of biological systems. In the filamentous fungus Neurospora crassa, multiple incompatibility genes mediate nonself recognition during vegetative growth One of these genes, un-24, encodes both nonself recognition function and the large subunit of a type I ribonucleotide reductase, an evolutionarily conserved enzyme that is essential for the conversion of NDP precursors into dNDPs for use in DNA synthesis. Nonself recognition systems are ubiquitous in diverse organisms as exemplified by restriction endonucleases in bacteria [1] and the major histocompatibility complex in vertebrates [2] In filamentous fungi, such as Neurospora crassa, nonself recognition occurs in both the sexual and vegetative phases [3]. This dual function of un-24 is of particular interest since it implicates a potential connection between DNA synthesis and nonself recognition-associated cell death
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