Biology subjects

Span, I.

Publications and source records attributed to Span, I..

3 recordsLinked to original sources

bioRxiv2026

Species Context Reverses PPIP5K Control of Fungal Morphogenesis and Actin Organization

Inositol pyrophosphates are conserved signaling molecules synthesized by the bifunctional PPIP5K enzymes, but how their cellular functions diversify across species remains poorly understood. Here, we compared the PPIP5K enzyme Asp1 in the fission yeasts Schizosaccharomyces pombe and Schizosaccharomyces japonicus and in the distantly related fungus Ustilago maydis. All three homologs retained a conserved kinase-phosphatase architecture and catalytic activity. However, whereas Asp1 produced broadly similar effects on actin organization and morphogenesis in S. pombe and U. maydis, its regulatory output was reversed in S. japonicus. In S. pombe and U. maydis, Asp1 positively supported Arp2/3-dependent actin functions, as loss of Asp1 increased sensitivity to the Arp2/3 inhibitor CK666. In contrast, deletion of asp1 in S. japonicus conferred strong CK666 resistance and caused excessive, spatially deregulated actin-patch organization. This opposing cytoskeletal phenotype was mirrored at the level of morphogenesis: Asp1 restricted the yeast-to-hypha transition in S. japonicus, whereas Asp1 was required for pseudohyphal growth in S. pombe and for filamentous development in U. maydis. However, the negative regulatory activity observed in S. japonicus was not an intrinsic property of the SjAsp1 protein. When expressed in S. pombe, SjAsp1 promoted invasive pseudohyphal growth, reproducing the regulatory output of the S. pombe Asp1 morphogenesis pathway rather than that of its native species. Similarly, SjAsp1 supported Arp2/3 functions when expressed in S. pombe. Thus, SjAsp1 adopted the functional behavior imposed by the host cellular environment. S. pombe Asp1 was originally identified as a suppressor of Arp2/3-complex mutant phenotypes, establishing a genetic connection between Asp1 and the actin nucleator. Extending this link, affinity enrichment with inositol pyrophosphates reagents recovered all seven subunits of the S. pombe Arp2/3 complex, providing biochemical support for a potential association between inositol pyrophosphate and Arp2/3. Together, these findings identify S. japonicus as a functional outlier in which a conserved PPIP5K pathway produces an opposing biological output. They further demonstrate that this divergence is determined primarily by species-specific cellular networks rather than by intrinsic differences in the Asp1 protein.

cell biology↗

bioRxiv2026

Molecular crowding: impacts on the activity of the 10-23 DNAzyme

The growing number of approved nucleic acid therapeutics illustrates the potential to treat diseases by targeting their genetic blueprints in vivo. The 10-23 DNAzyme is capable of cleaving a wide range of target RNA with high selectivity. However, its poor performance in vivo restricts its therapeutic application as gene silencing agent. Studies on ribozymes have shown that the crowded environment in cells and associated effects can impact ribozyme folding and thermostability, resulting in a change in activity. This opens up the question whether DNAzymes are also affected by molecular crowding. Here, we investigate the functional and structural influence of molecular crowding conditions on the 10-23 DNAzyme. The stability and activity of a PrP-specific 10-23 DNAzyme were examined in presence of PEG, dextran, and osmolytes. Our results indicate that osmolytes decrease DNAzyme activity in a concentration-dependent manner, while certain PEG and dextran concentrations promote activity. To rationalize our observations, we studied the cosolutes effect on physicochemical solution properties and the structure of the DNAzyme:RNA complex using FCS and SAXS. The data reveal that enhanced activity is observed under conditions where a combination of physiochemical properties matches an optimum that seems to be dependent on the metal ion cofactor. Structural influence under such conditions is indicated less. We propose that a certain degree of molecular crowding is required to favor a state, which allows for higher catalytic turnover. In addition, we show that the requirement for magnesium and manganese as a cofactor remains unchanged under the conditions applied. Our work contributes to a better understanding of how the cellular environment affects DNAzyme structure and function.

biophysics↗

bioRxiv2021

Iron-sulfur clusters are involved in post-translational arginylation

Eukaryotic arginylation is an essential post-translational modification that both modulates protein stability and regulates protein half-life through the N-degron pathway. Arginylation is catalyzed by a family of enzymes known as the arginyl-tRNA transferases (ATE1s), which are conserved across the eukaryotic domain. Despite its conservation and importance, little is known regarding the structure, mechanism, and regulation of ATE1s. In this work, we have discovered that ATE1s bind a previously unknown iron-sulfur cluster that is conserved across evolution. We have extensively characterized the nature of this iron-sulfur cluster, and we show that the presence of the iron-sulfur cluster is linked to alterations in arginylation efficacy. Finally, we demonstrate that the ATE1 iron-sulfur cluster is oxygen sensitive, which could be a molecular mechanism of the N-degron pathway to sense oxidative stress. Thus, our data provide the framework of a cluster-based paradigm of ATE1 regulatory control.

biochemistry↗