Biology subjects

Milo, S.

Publications and source records attributed to Milo, S..

4 recordsLinked to original sources

bioRxiv2026

UV-induced translation capacity is regulated developmentally in fungi.

In fast-growing cells, most RNA molecules encode ribosomal subunits. During growth arrest caused by DNA damage, ribosome biogenesis is reduced because it is a highly energy-demanding process. Here, we report that, in two filamentous fungi, UV radiation significantly induces ribosome levels and translation capacity. This UV-induced response is developmentally regulated and occurs after filament formation. Translation induction modestly but significantly enhances both UV survival and repair. While some of the mRNAs that are differentially associated with ribosomes following UV exposure are associated with DNA repair, most of them belong to modules involved in protein folding and gene expression. TOR signaling is required for the immediate UV induction of ribosome biogenesis genes. However, we provide evidence that UV induces mechanisms that operate either upstream of or in parallel with TOR. These findings suggest a novel type of response to DNA damage, in which the primary aim is to protect the integrity of gene expression from the detrimental effects of UV radiation on transcription.

microbiology↗

bioRxiv2026

The Ancient Origin and Dynamic Diversification of the Fungal Poly(ADP)-ribose Polymerase Protein Family

Poly(ADP-ribose) polymerases (PARPs) catalyze ADP-ribosylation, a conserved post- translational modification involved in DNA repair, transcriptional regulation, and chromatin remodeling. Although extensively studied in animals, the evolution and diversification of PARPs across the fungal kingdom remain largely unexplored. Here, we present the first kingdom-wide comparative genomic analysis of PARP proteins across 534 fungal species spanning eight phyla. We identified two primary conserved fungal PARP protein types corresponding to human PARP1 and PARP6. Both exhibit highly dynamic evolutionary histories characterized by frequent independent gain and loss events. Ancestral state reconstruction supports the presence of PARP1 in the last common ancestor of fungi, whereas the PARP6-like family has undergone repeated lineage-specific gains and losses. Fungal PARP6-like proteins retain a compact PARP catalytic domain fused to a C-terminal E2 ubiquitin-conjugating domain, whereas the PARP1 family displays extensive structural diversification through domain shuffling and lineage-specific fusions associated with DNA metabolism, chromatin remodeling, and signal transduction. Reconstruction of ancestral catalytic motifs across fungi and other eukaryotes revealed convergent evolution of a non-canonical H-Y-Y catalytic triad, with multiple motif variants co-occurring within individual proteins, suggesting functional diversification. In the Fusarium oxysporum species complex, we identified a lineage-specific expansion of the PARP family, driven exclusively by accessory chromosomes. Genomes with expanded PARP1 repertoires exhibited elevated basal PARylation, increased resistance to DNA-damaging agents that induce single strand breaks, and DNA damage- induced expression of accessory Parp genes. These findings reveal fungal PARPs as evolutionarily dynamic proteins that likely contribute to genome stability, adaptation, and pathogenicity.

evolutionary biology↗

bioRxiv2024

Repurposing CRISPR-Cas13 systems for robust mRNA trans-splicing

Type VI CRISPR enzymes have been developed as programmable RNA-guided Cas proteins for eukaryotic RNA editing. Notably, Cas13 has been utilized for site-targeted single base edits, demethylation, RNA cleavage or knockdown and alternative splicing. However, the ability to edit large stretches of mRNA transcripts remains a significant challenge. Here, we demonstrate that CRISPR-Cas13 systems can be repurposed to assist trans-splicing of exogenous RNA fragments into an endogenous pre-mRNA transcript, a method termed CRISPR Assisted mRNA Fragment Trans-splicing (CRAFT). Using split reporter-based assays, we evaluate orthogonal Cas13 systems, optimize guide RNA length and screen for optimal trans-splicing site(s) across a range of intronic targets. We achieve markedly improved editing of large 5 and 3 segments in different endogenous mRNAs across various mammalian cell types compared to other spliceosome-mediated trans-splicing methods. CRAFT can serve as a versatile platform for attachment of protein tags, studying the impact of multiple mutations/single nucleotide polymorphisms, modification of untranslated regions (UTRs) or replacing large segments of mRNA transcripts.

bioengineering↗

bioRxiv2019

The response to the DNA damaging agent methyl methanesulfonate in a fungal plant pathogen

DNA damage can cause mutations that in fungal plant pathogens lead to hypervirulence and resistance to pesticides. Almost nothing is known about the response of these fungi to DNA damage. We performed transcriptomic and phosphoproteomic analyses of Fusarium oxysporum exposed to a DNA alkylating agent (MMS). At the RNA level we observe massive induction of DNA repair pathways; the most surprising is the global genome nucleotide excision repair. Cul3, Cul4, several Ubiquitin-like ligases and other components of the protein degradation machinery are significantly induced. In agreement, we observed drug synergism between a proteasome inhibitor and MMS. While our data suggest that Yap1 and Xbp1 networks are similarly activated in response to damage in yeast and Fusarium we were able to observe Fusarium specific MMS-responsive modules. These include transcription/splicing modules that are upregulated and respiration that is down-regulated. In agreement, MMS treated cells are much more sensitive to a respiration inhibitor. At the phosphoproteomic level, Adenylate cyclase, which generates cAMP, is phosphorylated by MMS and forms a network of phosphorylated proteins that include cell cycle regulators and several MAPK pathways. Recently, there are many attempts to re-sequence isolates of fungal plant pathogens. These attempts often reveal changes in the genomes that may be explained by DNA damage exposure. Our analysis provides an important starting point in understanding how these genomic changes occur.

microbiology↗