No abstract

Phytoalexin glyceollins are soybean-specific antimicrobial compounds that are derived from the isoflavonoid pathway. They are synthesized by soybean in response to extrinsic stress such as pathogen attack or injury, thereby conferring partial resistance if synthesized rapidly at the site of infection and at the required concentration. Soybean produces multiple forms of glyceollins that result from the differential prenylation reaction catalyzed by prenyltransferases (PTs) on either the C-2 or C-4 carbon of a pterocarpan glycinol. The soybean genome contains 77 PT-encoding genes (GmPTs) where at least 11 are (iso)flavonoid-specific. Transcript accumulation of five candidates GmPTs was increased in response to Phytophthora sojae infection, suggesting their role in phytoalexin synthesis. The induced GmPTs localize to plastids and display tissue-specific expression. We have in this study identified two additional GmPTs: an isoflavone dimethylallyltransferase 3 (IDT3); and a glycinol 2-dimethylallyl transferase GmPT01. GmPT01 prenylates (-)-glycinol at the C-2 position, localizes in the plastid, and exhibits root-specific gene expression. Furthermore, its expression is induced rapidly in response to stress, and is associated with a quantitative trait loci linked with resistance to P. sojae. Based on these results, we conclude that GmPT01 are possibly one of the loci involved in conferring partial resistance against stem and root rot disease in soybean.

Candida albicans is a microbial fungus that exists as a commensal member of the human microbiome and an opportunistic pathogen. Cell surface-associated adhesin proteins play a crucial role in C. albicans’ ability to undergo cellular morphogenesis, develop robust biofilms, colonize, and cause infection in a host. However, a comprehensive analysis of the role and relationships between these adhesins has not been explored. We previously established a CRISPR-based platform for efficient generation of single- and double-gene deletions in C. albicans, which was used to construct a library of 144 mutants, comprising 12 unique adhesin genes deleted singly, and every possible combination of double deletions. Here, we exploit this adhesin mutant library to explore the role of adhesin proteins in C. albicans virulence. We perform a comprehensive, high-throughput screen of this library, using Caenorhabditis elegans as a simplified model host system, which identified mutants critical for virulence and significant genetic interactions. We perform follow-up analysis to assess the ability of high- and low-virulence strains to undergo cellular morphogenesis and form biofilms in vitro, as well as to colonize the C. elegans host. We further perform genetic interaction analysis to identify novel significant negative genetic interactions between adhesin mutants, whereby combinatorial perturbation of these genes significantly impairs virulence, more than expected based on virulence of the single mutant constituent strains. Together, this study yields important new insight into the role of adhesins, singly and in combinations, in mediating diverse facets of virulence of this critical fungal pathogen.

Nutrient adaptation is key in limiting environments for the promotion of microbial growth and survival. In microbial systems, iron is an essential component for many cellular processes, and bioavailability varies greatly among different conditions. In the bacterium, Klebsiella pneumoniae, the impact of iron limitation is known to alter transcriptional expression of iron-acquisition pathways and influence secretion of ironbinding siderophores, however, a comprehensive view of iron limitation at the protein level remains to be defined. Here, we apply a mass-spectrometry-based quantitative proteomics strategy to profile the global impact of iron limitation on the cellular proteome and extracellular environment (secretome) of K. pneumoniae. Our data define the impact of iron on proteins involved in transcriptional regulation and emphasize the modulation of a vast array of proteins associated with iron acquisition, transport, and binding. We also identify proteins in the extracellular environment associated with conventional and non-conventional modes of secretion, as well as vesicle release. In particular, we demonstrate a new role for Lon protease in promoting iron homeostasis outside of the cell. Characterization of a Lon protease mutant in K. pneumoniae validates roles in bacterial growth, cell division, and virulence, and uncovers novel degradation candidates of Lon protease associated with improved iron utilization strategies in the absence of the enzyme. Overall, we provide evidence of unique connections between Lon and iron in a bacterial system and suggest a new role for Lon protease in the extracellular environment during nutrient limitation.