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Li, X.-Z.

Publications and source records attributed to Li, X.-Z..

7 recordsLinked to original sources
bioRxiv2026

Differential pairings of MAT1-1-1 and MAT1-2-1 protein variants with heteromorphic tertiary structures produced by Ophiocordyceps sinensis strains

The MAT_HMGbox domain in the MAT1-1-1 protein and the HMG-box_ROX1-like domain in the MAT1-2-1 protein play essential roles in DNA binding and regulating the transcription of genes that control sexual reproduction in Ophiocordyceps sinensis. Previous studies have reported differential occurrences, differential transcription, and alternative splicing of the MAT1-1-1, MAT1-2-1, and pheromone receptor genes in Hirsutella sinensis (Genotype #1 among 17 genome-independent genotypes of O. sinensis fungi). In this study, we further demonstrated that the DNA-binding domains of the paired MAT1-1-1 and MAT1-2-1 proteins derived from each of the 20 purportedly "pure" O. sinensis strains exhibit heteromorphic tertiary structures, as predicted by AlphaFold 3D structural modeling. These differentially paired mating proteins are characterized by different truncations, diverse 1-4 amino acid substitutions at distinct sites, altered hydrophobicity and secondary structures, and heteromorphic tertiary structures, indicating divergency in the fungal origins of the mating proteins within the mycologically and genetically impure O. sinensis strains. Our findings support the self-sterility hypothesis for O. sinensis and likely ensure the fidelity and genetic diversity of heterothallic or hybrid reproduction during the lifecycle of the Cordyceps sinensis insect-fungal complex.

biochemistry↗
bioRxiv2025

Altered stereostructures of the DNA binding domains in the mutant mating proteins of Ophiocordyceps sinensis and the Cordyceps sinensis insect-fungal complex

The MAT_HMGbox and HMG-box_ROX1-like domains of the MAT1-1-1 and MAT1-2-1 proteins, respectively, play essential roles in DNA binding and the downstream regulation of gene transcription that controls sexual reproduction in Ophiocordyceps sinensis. Alternative splicing, differential occurrence and transcription of the MAT1-1-1 and MAT1-2-1 genes have been revealed in Hirsutella sinensis (Genotype #1 of the 17 O. sinensis genotypes), suggesting the occurrence of self-sterility under heterothallic or hybrid outcrossing. This study demonstrates that the MAT_HMGbox domains of MAT1-1-1 proteins in wild-type Cordyceps sinensis isolates cluster into 5 clades with or without branches in the Bayesian clustering tree and belong to diverse 3D structural morphs under 19 AlphaFold codes. The HMG-box_ROX1-like domains of MAT1-2-1 proteins cluster into 2 clades with branches in the Bayesian clustering tree and belong to diverse 3D structural morphs under 25 AlphaFold codes. Correlation analysis reveals that 1-3 amino acid substitutions at various sites in the DNA-binding domains of the mutant mating proteins result in alterations in the hydrophobicity and the secondary and tertiary structures of the DNA-binding domains of the proteins. Fungal origin analysis reveals heterospecific fungal sources or genome-independent, genotypic sources of the mating proteins with stereostructure variations in the wild-type C. sinensis isolates that represent the changed functionalities of the proteins in regulating and ensuring the accuracy and genetic diversity of the heterothallic and hybrid sexual reproductive processes of O. sinensis during the lifecycle of the C. sinensis insect-fungi complex.

biochemistry↗
bioRxiv2022

Multiple GC-biased repetitive ITS copies in the Hirsutella sinensis genome are not generated by RIP mutagenesis involving transition point mutations

It has been hypothesized that AT-biased genotypes of Ophiocordyceps sinensis are generated through repeat-induced point mutation (RIP) and coexist as permanently nonfunctional internal transcribed spacer (ITS) pseudogenes in the genome of Hirsutella sinensis (GC-biased Genotype #1 of O. sinensis). This study examined the H. sinensis genome, which contains multiple repetitive ITS copies (GC content: 64.7{+/-}0.33%) with multiple insertion/deletion and transversion alleles, which were not generated through RIP mutagenesis that theoretically causes cytosine-to-thymine (C-to-T) and guanine-to-adenine (G-to-A) transitions. The repetitive ITS copies in the H. sinensis genome were found to be genetically and phylogenetically distinct from the AT-biased O. sinensis genotypes (GC content: 51.1{+/-}1.69%), which possess multiple transition alleles. The sequences of Genotypes #2-17, both GC- and AT-biased, are absent from the H. sinensis genome; these genotypes belong to interindividual O. sinensis fungi and differentially occur in different compartments of natural Cordyceps sinensis, with dynamic alterations in abundance occurring in an asynchronous, disproportional manner during C. sinensis maturation. Metatranscriptomic analyses of natural C. sinensis revealed the transcriptional silencing of 5.8S genes in all C. sinensis- colonizing fungi, including H. sinensis. The transcription assay reported by Li et al. [1] provided unsound, controversial evidence indicating that the 5.8S genes of AT-biased genotypes are nonfunctional pseudogenes. In addition to the single ITS locus analysis, repetitive genomic copies were also examined at multiple loci in the H. sinensis genome, and approximately 8.2% of the authentic genes had repetitive copies, including various transitions, transversions, and insertions/deletions. The transcripts for the repetitive copies, regardless of the decreases, increases, or bidirectional changes in the AT content, were identified in the H. sinensis transcriptome. These results are inconsistent with those of RIP mutagenesis, which generates pseudogenic, nonfunctional, repetitive copies. In conclusion, AT-biased genotypes of O. sinensis might have evolved through evolutionary mechanisms from a common ancestor over the long course of evolution, in parallel with GC-biased Genotype #1 H. sinensis.

molecular biology↗
bioRxiv2022

Altered GC- and AT-biased genotypes of Ophiocordyceps sinensis in the stromal fertile portions and ascospores of natural Cordyceps sinensis

ObjectiveTo examine multiple genotypes of Ophiocordyceps sinensis in a semi-quantitative manner in the stromal fertile portion (SFP) densely covered with numerous ascocarps and ascospores of natural Cordyceps sinensis and to outline the dynamic alterations of the coexisting O. sinensis genotypes in different developmental phases. MethodsMature Cordyceps sinensis specimens were harvested and continuously cultivated in our laboratory (altitude 2,254 m). The SFPs (with ascocarps) and fully and semi-ejected ascospores were collected for histological and molecular examinations. Biochip-based single nucleotide polymorphism (SNP) MALDI-TOF mass spectrometry (MS) was used to genotype multiple O. sinensis mutants in the SFPs and ascospores. ResultsMicroscopic analysis revealed distinct morphologies of the SFPs (with ascocarps) before and after ascospore ejection and SFP of developmental failure, which, along with the fully and semi-ejected ascospores, were subjected to SNP MS genotyping analysis. Mass spectra showed the coexistence of GC- and AT-biased genotypes of O. sinensis that were genetically and phylogenetically distinct in the SFPs before and after ejection and of developmental failure and in fully and semi-ejected ascospores. The intensity ratios of MS peaks were dynamically altered in the SFPs and the fully and semi-ejected ascospores. Mass spectra also showed transversion mutation alleles of unknown upstream and downstream sequences with altered intensities in the SFPs and ascospores. Genotype #5 of AT-biased Cluster-A maintained a high intensity in all SFPs and ascospores. An MS peak with a high intensity containing AT-biased Genotypes #6 and #15 in pre-ejection SFPs was significantly attenuated after ascospore ejection. The abundance of Genotypes #5-6 and #16 of AT-biased Cluster-A was differentially altered in the fully and semi-ejected ascospores that were collected from the same Cordyceps sinensis specimens. ConclusionMultiple O. sinensis genotypes coexisted in different combinations with altered abundances in the SFPs prior to and after ejection, the SFP of developmental failure, and the two types of ascospores of Cordyceps sinensis, demonstrating their genomic independence. Metagenomic fungal members present in different combinations and with dynamic alterations play symbiotic roles in different compartments of natural Cordyceps sinensis.

molecular biology↗
bioRxiv2022

Differential coexistence of multiple genotypes of Ophiocordyceps sinensis in the stromata, ascocarps and ascospores of natural Cordyceps sinensis

ObjectiveTo examine the differential occurrence of Ophiocordyceps sinensis genotypes in the stroma, stromal fertile portion (SFP) densely covered with numerous ascocarps, and ascospores of natural Cordyceps sinensis. MethodsImmature and mature C. sinensis specimens were harvested. Mature C. sinensis specimens were continuously cultivated in our laboratory (altitude 2,200 m). The SFPs (with ascocarps) and ascospores of C. sinensis were collected for microscopic and molecular analyses using species-/genotype-specific primers. Sequences of mutant genotypes of O. sinensis were aligned with that of Genotype #1 Hirsutella sinensis and compared phylogenetically using a Bayesian majority-rule method. ResultsFully and semiejected ascospores were collected from the same specimens. The semiejected ascospores tightly adhered to the surface of the asci as observed by the naked eye and under optical and confocal microscopies. The multicellular heterokaryotic ascospores showed uneven staining of nuclei. The immature and mature stromata, SFPs (with ascocarps) and ascospores were found to differentially contain several GC- and AT-biased genotypes of O. sinensis, Samsoniella hepiali, and an AB067719-type fungus. The genotypes within AT-biased Cluster-A in the Bayesian tree occurred in all compartments of C. sinensis, but those within AT-biased Cluster-B were present in immature and mature stromata and SPFs but absent in the ascospores. Genotype #13 of O. sinensis was present in semi-ejected ascospores and Genotype #14 in fully ejected ascospores. GC-biased Genotypes #13-14 featured large DNA segment substitutions and genetic material recombination between the genomes of the parental fungi (H. sinensis and the AB067719-type fungus). These ascosporic offspring genotypes combined with varying abundances of S. hepiali in the 2 types of ascospores participated in the control of the development, maturation and ejection of the ascospores. ConclusionMultiple genotypes of O. sinensis coexist differentially in the stromata, SFPs and 2 types of C. sinensis ascospores, along with S. hepiali and the AB067719-type fungus. The fungal components in different combinations and their dynamic alterations in the compartments of C. sinensis during maturation play symbiotic roles in the lifecycle of natural C. sinensis.

molecular biology↗
bioRxiv2019

Chemistry of cation hydration and conduction in a skeletal muscle ryanodine receptor

Ryanodine receptors (RyRs) are Ca2+-regulated Ca2+ channels of 2.2-megadalton in muscles and neurons for calcium signaling. How Ca2+ regulates ion conduction in the RyR channels remains elusive. We determined a 2.6-[A] cryo-EM structure of rabbit skeletal muscle RyR1, and used multiscale dynamics simulations to elucidate cation interactions with RyR1. We investigated 21 potential cation-binding sites that may together rationalize biphasic Ca2+ response of RyR1. The selectivity filter captures a cation hydration complex by hydrogen-bonding with both the inner and outer hydration shells of water molecules. Molecular dynamics simulations suggest that adjacent Ca2+ ions moving in concert along ion-permeation pathway are separated by at least two cation-binding sites. Our analysis reveals that RyR1 has been evolved to favor its interactions with two hydration shells of cations.

biochemistry↗