Yuanxi Xiao

Bio: Yuanxi Xiao is an academic researcher from Shandong University. The author has contributed to research in topics: Mutant & Wild type. The author has an hindex of 3, co-authored 3 publications receiving 15 citations.

The Lonely Guy (LOG) Homologue SiRe_0427 from the Thermophilic Archaeon Sulfolobus islandicus REY15A Is a Phosphoribohydrolase Representing a Novel Group.

Abstract: Lonely Guy (LOG) proteins are important enzymes in cellular organisms, which catalyze the final step in the production of biologically active cytokinins via dephosphoribosylation. LOG proteins are vital enzymes in plants for the activation of cytokinin precursors, which is crucial for plant growth and development. In fungi and bacteria, LOGs are implicated in pathogenic or nonpathogenic interactions with their plant hosts. However, LOGs have also been identified in the human pathogen Mycobacterium tuberculosis, and the accumulation of cytokinin-degraded products, aldehydes, within bacterial cells is lethal to the bacterium in the presence of nitric oxide, suggesting diverse roles of LOGs in various species. In this study, we conducted biochemical and genetic analysis of a LOG homologue, SiRe_0427, from the hyperthermophilic archaeon Sulfolobus islandicus REY15A. The protein possessed the LOG motif GGGxGTxxE and exhibited phosphoribohydrolase activity on adenosine-5-monophosphate (AMP), similar to LOGs from eukaryotes and bacteria. Alanine mutants at either catalytic residues or substrate binding sites lost their activity, resembling other known LOGs. SiRe_0427 is probably a homotetramer, as revealed by size exclusion chromatography and chemical cross-linking. We found that the gene encoding SiRe_0427 could be knocked out; however, the Δsire_0427 strain exhibited no apparent difference in growth compared to the wild type, nor did it show any difference in sensitivity to UV irradiation under our laboratory growth conditions. Overall, these findings indicate that archaeal LOG homologue is active as a phosphoribohydrolase. IMPORTANCE Lonely Guy (LOG) is an essential enzyme for the final biosynthesis of cytokinins, which regulate almost every aspect of growth and development in plants. LOG protein was originally discovered 12 years ago in a strain of Oryza sativa with a distinct floral phenotype of a single stamen. Recently, the presence of LOG homologues has been reported in Mycobacterium tuberculosis, an obligate human pathogen. To date, active LOG proteins have been reported in plants, pathogenic and nonpathogenic fungi, and bacteria, but there have been no experimental reports of LOG protein from archaea. In the current work, we report the identification of a LOG homologue active on AMP from Sulfolobus islandicus REY15A, a thermophilic archaeon. The protein likely forms a tetramer in solution and represents a novel evolutionary lineage. The results presented here expand our knowledge regarding proteins with phosphoribohydrolase activities and open an avenue for studying signal transduction networks of archaea and potential applications of LOG enzymes in agriculture and industry.

Functional Analysis of the NucS/EndoMS of the Hyperthermophilic Archaeon Sulfolobus islandicus REY15A.

Abstract: EndoMS is a recently identified mismatch specific endonuclease in Thermococcales of Archaea and Mycobacteria of Bacteria. The homologs of EndoMS are conserved in Archaea and Actinobacteria, where classic MutS-MutL-mediated DNA mismatch repair pathway is absent or non-functional. Here, we report a study on the in vitro mismatch cleavage activity and in vivo function of an EndoMS homolog (SisEndoMS) from Sulfolobus islandicus REY15A, the model archaeon belonging to Crenarchaeota. SisEndoMS is highly active on duplex DNA containing G/T, G/G, and T/T mismatches. Interestingly, the cleavage activity of SisEndoMS is stimulated by the heterotrimeric PCNAs, and when Mn2+ was used as the co-factor instead of Mg2+, SisEndoMS was also active on DNA substrates containing C/T or A/G mismatches, suggesting that the endonuclease activity can be regulated by ion co-factors and accessory proteins. We compared the spontaneous mutation rate of the wild type strain REY15A and ∆endoMS by counter selection against 5-fluoroorotic acid (5-FOA). The endoMS knockout mutant had much higher spontaneous mutation rate (5.06 × 10-3) than that of the wild type (4.6 × 10-6). A mutation accumulation analysis also showed that the deletion mutant had a higher mutation occurrence than the wild type, with transition mutation being the dominant, suggesting that SisEndoMS is responsible for mutation avoidance in this hyperthermophilic archaeon. Overexpression of the wild type SisEndoMS in S. islandicus resulted in retarded growth and abnormal cell morphology, similar to strains overexpressing Hje and Hjc, the Holliday junction endonucleases. Transcriptomic analysis revealed that SisEndoMS overexpression led to upregulation of distinct gene including the CRISPR-Cas IIIB system, methyltransferases, and glycosyltransferases, which are mainly localized to specific regions in the chromosome. Collectively, our results support that EndoMS proteins represent a noncanonical DNA repair pathway in Archaea. The mechanism of the mismatch repair pathway in Sulfolobus which have a single chromosome is discussed.

The Hulks and the Deadpools of the Cytokinin Universe: A Dual Strategy for Cytokinin Production, Translocation, and Signal Transduction.

Abstract: Cytokinins are plant hormones, derivatives of adenine with a side chain at the N6-position. They are involved in many physiological processes. While the metabolism of trans-zeatin and isopentenyladenine, which are considered to be highly active cytokinins, has been extensively studied, there are others with less obvious functions, such as cis-zeatin, dihydrozeatin, and aromatic cytokinins, which have been comparatively neglected. To help explain this duality, we present a novel hypothesis metaphorically comparing various cytokinin forms, enzymes of CK metabolism, and their signalling and transporter functions to the comics superheroes Hulk and Deadpool. Hulk is a powerful but short-lived creation, whilst Deadpool presents a more subtle and enduring force. With this dual framework in mind, this review compares different cytokinin metabolites, and their biosynthesis, translocation, and sensing to illustrate the different mechanisms behind the two CK strategies. This is put together and applied to a plant developmental scale and, beyond plants, to interactions with organisms of other kingdoms, to highlight where future study can benefit the understanding of plant fitness and productivity.

The Origins and Roles of Methylthiolated Cytokinins: Evidence From Among Life Kingdoms.

Abstract: Cytokinins (CKs) are a group of adenine-derived, small signaling molecules of crucial importance for growth and multiple developmental processes in plants. Biological roles of classical CKs: isopentenyladenine (iP), trans and cis isomers of zeatin (tZ, cZ), and dihydrozeatin, have been studied extensively and their functions are well defined in many aspects of plant physiology. In parallel, extensive knowledge exists for genes involved in tRNA modifications that lead to the production of tRNA-bound methylthiolated CKs, especially in bacterial and mammalian systems. However, not much is known about the origins, fates, and possible functions of the unbound methylthiolated CKs (2MeS-CKs) in biological systems. 2MeS-CKs are the free base or riboside derivatives of iP or Z-type CKs, modified by the addition of a thiol group (-SH) at position 2 of the adenine ring that is subsequently methylated. Based on the evidence to date, these distinctive CK conjugates are derived exclusively via the tRNA degradation pathway. This review summarizes the knowledge on the probable steps involved in the biosynthesis of unbound 2MeS-CKs across diverse kingdoms of life. Furthermore, it provides examples of CK profiles of organisms from which the presence of 2MeS-CKs have been detected and confirms a close association and balance between the production of classical CKs and 2MeS-CKs. Finally, it discusses available reports regarding the possible physiological functions of 2MeS-CKs in different biological systems.

Biochemical and Structural Aspects of Cytokinin Biosynthesis and Degradation in Bacteria

Abstract: It has been known for quite some time that cytokinins, hormones typical of plants, are also produced and metabolized in bacteria. Most bacteria can only form the tRNA-bound cytokinins, but there are examples of plant-associated bacteria, both pathogenic and beneficial, that actively synthesize cytokinins to interact with their host. Similar to plants, bacteria produce diverse cytokinin metabolites, employing corresponding metabolic pathways. The identification of genes encoding the enzymes involved in cytokinin biosynthesis and metabolism facilitated their detailed characterization based on both classical enzyme assays and structural approaches. This review summarizes the present knowledge on key enzymes involved in cytokinin biosynthesis, modifications, and degradation in bacteria, and discusses their catalytic properties in relation to the presence of specific amino acid residues and protein structure.

Unlocking the potential of insect and ruminant host symbionts for recycling of lignocellulosic carbon with a biorefinery approach: a review.

Abstract: Uprising fossil fuel depletion and deterioration of ecological reserves supply have led to the search for alternative renewable and sustainable energy sources and chemicals. Although first generation biorefinery is quite successful commercially in generating bulk of biofuels globally, the food versus fuel debate has necessitated the use of non-edible feedstocks, majorly waste biomass, for second generation production of biofuels and chemicals. A diverse class of microbes and enzymes are being exploited for biofuels production for a series of treatment process, however, the conversion efficiency of wide range of lignocellulosic biomass (LCB) and consolidated way of processing remains challenging. There were lot of research efforts in the past decade to scour for potential microbial candidate. In this context, evolution has developed the gut microbiota of several insects and ruminants that are potential LCB degraders host eco-system to overcome its host nutritional constraints, where LCB processed by microbiomes pretends to be a promising candidate. Synergistic microbial symbionts could make a significant contribution towards recycling the renewable carbon from distinctly abundant recalcitrant LCB. Several studies have assessed the bioprospection of innumerable gut symbionts and their lignocellulolytic enzymes for LCB degradation. Though, some reviews exist on molecular characterization of gut microbes, but none of them has enlightened the microbial community design coupled with various LCB valorization which intensifies the microbial diversity in biofuels application. This review provides a deep insight into the significant breakthroughs attained in enrichment strategy of gut microbial community and its molecular characterization techniques which aids in understanding the holistic microbial community dynamics. Special emphasis is placed on gut microbial role in LCB depolymerization strategies to lignocellulolytic enzymes production and its functional metagenomic data mining eventually generating the sugar platform for biofuels and renewable chemicals production.