Mosaic fungal individuals have the potential to evolve within a single generation

Although cells of mushroom-producing fungi typically contain paired haploid nuclei (n + n), most Armillaria gallica vegetative cells are uninucleate. As vegetative nuclei are produced by fusions of paired haploid nuclei, they are thought to be diploid (2n). Here we report finding haploid vegetative nuclei in A. gallica at multiple sites in southeastern Massachusetts, USA. Sequencing multiple clones of a single-copy gene isolated from single hyphal filaments revealed nuclear heterogeneity both among and within hyphae. Cytoplasmic bridges connected hyphae in field-collected and cultured samples, and we propose nuclear migration through bridges maintains this nuclear heterogeneity. Growth studies demonstrate among- and within-hypha phenotypic variation for growth in response to gallic acid, a plant-produced antifungal compound. The existence of both genetic and phenotypic variation within vegetative hyphae suggests that fungal individuals have the potential to evolve within a single generation in response to environmental variation over time and space

The UVSSA protein is part of a genome integrity homeostasis network with links to transcription-coupled DNA repair and ATM signaling.

SignificanceTranscription-coupled repair (TCR) involves four core proteins: CSA, CSB, USP7, and UVSSA. CSA and CSB are mutated in the severe human neurocutaneous disease Cockayne syndrome. In contrast UVSSA is a mild photosensitive disease in which a mutated protein sequence prevents recruitment of USP7 protease to deubiquitinate and stabilize CSB. We deleted the UVSSA protein using CRISPR-Cas9 in an aneuploid cell line, HEK293, and determined the functional consequences. The knockout cell line was sensitive to transcription-blocking lesions but not sensitive to oxidative agents or PARP inhibitors, unlike CSB. Knockout of UVSSA also activated ATM, like CSB, in transcription-arrested cells. The phenotype of UVSSA, especially its rarity, suggests that many TCR-deficient patients and tumors fail to be recognized clinically

The UVSSA protein is part of a genome integrity homeostasis network with links to transcription-coupled DNA repair and ATM signaling

The UVSSA (KIAA1530) protein is a component of transcription-coupled repair which, together with the CSA(ERCC8) and CSB(ERCC6) proteins cooperates to relieve transcription-blocking DNA damage. Mutations in CSA and CSB are found in Cockayne syndrome (CS), which is a human recessively inherited photosensitive, neurocutaneous, aging disorder. Mutations in UVSSA, in contrast, are found in the rare mild photosensitive syndrome (UV(s)) that lacks the noncutaneous complications of CSA or CSB patients. In this study we deployed CRISPR to disrupt exon I of the UVSSA gene in the human embryonic kidney cell line HEK293. Elimination of the UVSSA protein was confirmed by Western blotting and the knockout cells displayed the predicted sensitivity to transcription blocking lesions caused by illudin, cisplatin, and ultraviolet light, just as in CS cell lines. Transcription arrest in a UVSSA knockout cell line resulted in ATM-dependent phosphorylation of H2Ax and delayed DNA synthesis, relieved by an inhibitor of ATM. Loss of UVSSA protein did not, however, increase sensitivity to oxidative damage or to inhibitors of poly (ADP)ribose polymerase, unlike reported in CSB cells. We discuss this in terms of the likely commutative interplay of factors in CS. We anticipate that this knockout cell line will advance understanding of this and possibly related transcription-coupled DNA repair diseases