Xie J et al. (2016) IEEE/ACM Trans Comput Biol Bioinform "An Adaptive Hybrid Algorithm for Global Network Alignment."

Xie J, Lei J, Tan J, Xiang C, Nie Q, Ma J, Wen T

[IEEE/ACM Trans Comput Biol Bioinform, 2016]

It is challenging to obtain reliable and optimal mapping between networks for alignment algorithms when both nodal and topological structures are taken into consideration due to the underlying NP-hard problem. Here, we introduce an adaptive hybrid algorithm that combines the classical Hungarian algorithm and the Greedy algorithm (HGA) for the global alignment of biomolecular networks. With this hybrid algorithm, every pair of nodes with one in each network is first aligned based on node information (e.g., their sequence attributes) and then followed by an adaptive and convergent iteration procedure for aligning the topological connections in the networks. For four well-studied protein interaction networks, i.e., C.elegans, yeast, D.melanogaster, and human, applications of HGA lead to improved alignments in acceptable running time. The mapping between yeast and human PINs obtained by the new algorithm has the largest value of common gene ontology (GO) terms compared to those obtained by other existing algorithms, while it still has lower Mean normalized entropy (MNE) and good performances on several other measures. Overall, the adaptive HGA is effective and capable of providing good mappings between aligned networks in which the biological properties of both the nodes and the connections are important.

Xie J et al. (2023) PLoS Genet "MBL-1 and EEL-1 affect the splicing and protein levels of MEC-3 to control dendrite complexity."

Wang X, Tugizova M, Shen K, Zou W, Xie J

[PLoS Genet, 2023]

Transcription factors (TFs) play critical roles in specifying many aspects of neuronal cell fate including dendritic morphology. How TFs are accurately regulated during neuronal morphogenesis is not fully understood. Here, we show that LIM homeodomain protein MEC-3, the key TF for C. elegans PVD dendrite morphogenesis, is regulated by both alternative splicing and an E3 ubiquitin ligase. The mec-3 gene generates several transcripts by alternative splicing. We find that mbl-1, the orthologue of the muscular dystrophy disease gene muscleblind-like (MBNL), is required for PVD dendrite arbor formation. Our data suggest mbl-1 regulates the alternative splicing of mec-3 to produce its long isoform. Deleting the long isoform of mec-3(deExon2) causes reduction of dendrite complexity. Through a genetic modifier screen, we find that mutation in the E3 ubiquitin ligase EEL-1 suppresses mbl-1 phenotype. eel-1 mutants also suppress mec-3(deExon2) mutant but not the mec-3 null phenotype. Loss of EEL-1 alone leads to excessive dendrite branches. Together, these results indicate that MEC-3 is fine-tuned by alternative splicing and the ubiquitin system to produce the optimal level of dendrite branches.

Xie J et al. (2019) Curr Biol "Regulation of the Elongation Phase of Protein Synthesis Enhances Translation Accuracy and Modulates Lifespan."

O'Keefe L, Jensen KB, Coldwell MJ, de Souza Alves V, Liu R, Lenchine RV, von der Haar T, Wang X, Proud CG, Xie J

[Curr Biol, 2019]

Maintaining accuracy during protein synthesis is crucial to avoid producing misfolded and/or non-functional proteins. The target of rapamycin complex 1 (TORC1) pathway and the activity of the protein synthesis machinery are known to negatively regulate lifespan in many organisms, although the precise mechanisms involved remain unclear. Mammalian TORC1 signaling accelerates the elongation stage of protein synthesis by inactivating eukaryotic elongation factor 2 kinase (eEF2K), which, when active, phosphorylates and inhibits eEF2, which mediates the movement of ribosomes along mRNAs, thereby slowing down the rate of elongation. We show that eEF2K enhances the accuracy of protein synthesis under a range of conditions and in several cell types. For example, our data reveal it links mammalian (m)TORC1 signaling to the accuracy of translation. Activation of eEF2K decreases misreading or termination readthrough errors during elongation, whereas knocking down or knocking out eEF2K increases their frequency. eEF2K also promotes the correct recognition of start codons in mRNAs. Reduced translational fidelity is known to correlate with shorter lifespan. Consistent with this, deletion of the eEF2K ortholog or other factors implicated intranslation fidelity in Caenorhabditis elegans decreases lifespan, and eEF2K is required for lifespan extension induced by nutrient restriction. Our data uncover a novel mechanism linking nutrient supply, mTORC1 signaling, and the elongation stage of protein synthesis, which enhances the accuracy of protein synthesis. Our data also indicate that modulating translation elongation and its fidelity affects lifespan.

Xie J et al. (2023) Food Funct "Astaxanthin reduces fat storage in a fat-6/fat-7 dependent manner determined using high fat Caenorhabditis elegans."

Xiao J, Xie J, Hou X, Cao Y, Liu X, He W

[Food Funct, 2023]

Although astaxanthin has been shown to have high potential for weight loss, the specific action site and signal pathway generally cannot be confirmed in other animal models. This prevents us from finding therapeutic targets. Hence, we further illuminated its efficacy and specific action sites by using <i>Caenorhabditis elegans</i> (<i>C. elegans</i>). In this study, 60 &#x3bc;M astaxanthin supplementation reduced overall fat deposition and triglyceride levels by 21.47% and 22.00% (<i>p</i> &lt; 0.01). The content of large lipid droplets was reversed after astaxanthin treatment, and the ratio of oleic acid/stearic acid (C18:1&#x394;9/C18:0) decreased significantly, which were essential substrates for triglyceride biosynthesis. In addition, astaxanthin prevented obesity caused by excessive energy accumulation and insufficient energy consumption. Furthermore, the above effects were induced by <i>sbp-1</i>/<i>mdt-15</i> and insulin/insulin-like growth factor pathways, and finally co-regulated the specific site-<i>fat-6</i> and <i>fat-7</i> down-regulation. These results provided insight into therapeutic targets for future astaxanthin as a nutritional health product to relieve obesity.

Xie J et al. (2005) J Neurochem "Mutagenesis analysis of the serotonin 5-HT2C receptor and a Caenorhabditis elegans 5-HT2 homologue: conserved residues of helix 4 and helix 7 contribute to agonist-dependent activation of 5-HT2 receptors."

Xie J, Dernovici S, Ribeiro P

[J Neurochem, 2005]

An alignment of serotonin [5-hydroxytryptamine (5-HT)] G protein-coupled receptors identified a lysine at position 4.45 (helix 4) and a small polar residue (serine or cysteine) at 7.45 (helix 7) that occur exclusively in the 5-HT2 receptor family. Other serotonin receptors have a hydrophobic amino acid, typically a methionine, at 4.45 and an invariant asparagine at 7.45. The functional significance of these class-specific substitutions was tested by site-directed mutagenesis of two distantly related 5-HT2 receptors, Caenorhabditis elegans 5-HT2ce and rat 5-HT2C. Residues 4.45 and 7.45 were each mutated to a methionine and asparagine, respectively, or an alanine and the resulting constructs were tested for activity. A K4.45M mutation decreased serotonin-dependent activity (Emax) of the rat 5-HT2C receptor by 60% and that of the C. elegans homologue by 40%, as determined by a fluorometric plate-based calcium assay. The rat mutant also exhibited nearly sixfold higher agonist binding affinity and significantly lower constitutive activity compared with wildtype. Mutagenesis of S7.45 in the C. elegans receptor increased serotonin binding affinity by up to 25-fold and decreased Emax by up to 65%. The same mutations of the cognate C7.45 in rat 5-HT2C produced a smaller fourfold change in the affinity for serotonin and decreased agonist efficacy by up to 50%. Substitutions of S/C7.45 did not produce a significant change in the basal activity of either receptor. All mutants tested exhibited levels of receptor expression similar to the corresponding wildtype based on measurements of specific [3H]-mesulergine binding or flow cytometry analyses. Taken together, these results suggest that K4.45 and S/C7.45 play an important role in the conformational rearrangements leading to agonist-induced activation of 5-HT2 receptors.

Xie S et al. (2022) J Cell Sci "The retromer complex regulates C. elegans development and mammalian ciliogenesis."

Williams A, Caplan S, Naslavsky N, Iyer J, Duran R, Davis A, Mathew D, Dierlam C, Xie S, Smith E

[J Cell Sci, 2022]

The mammalian retromer consists of subunits VPS26, VPS29 and VPS35, and a loosely-associated sorting nexin (SNX) heterodimer or a variety of other SNX proteins. Despite involvement in yeast and mammalian cell trafficking, retromer's role in development is poorly understood, and its impact on primary ciliogenesis remains unknown. Using CRISPR-Cas9 editing, we demonstrate that vps-26 knockout worms have reduced brood sizes, impaired vulval development, and decreased body length, which have been linked to ciliogenesis defects. While preliminary studies did not identify worm ciliary defects, and impaired development limited additional ciliogenesis studies, we turned to mammalian cells to investigate the role of retromer in ciliogenesis. VPS35 localized to the primary cilium of mammalian cells, and depletion of VPS26, VPS35, VPS29, SNX1, SNX2, SNX5 or SNX27 led to decreased ciliogenesis. Retromer also coimmunoprecipitated with the centriolar protein, CP110, and was required for its removal from the mother centriole. Herein, we characterize new roles for the retromer in C. elegans development and in the regulation of ciliogenesis in mammalian cells, suggesting a novel role for the retromer in CP110 removal from the mother centriole.

Xie C et al. (2024) EMBO J "Neurons dispose of hyperactive kinesin into glial cells for clearance."

Chen Z, Chen G, Feng W, Xie C, Mohamed MA, Sonar P, Wang Y, Cleetus A, Li M, Ou G, Huang P, Oekten Z, Lei K, Li D

[EMBO J, 2024]

Microtubule-based kinesin motor proteins are crucial for intracellular transport, but their hyperactivation can be detrimental for cellular functions. This study investigated the impact of a constitutively active ciliary kinesin mutant, OSM-3CA, on sensory cilia in C. elegans. Surprisingly, we found that OSM-3CA was absent from cilia but underwent disposal through membrane abscission at the tips of aberrant neurites. Neighboring glial cells engulf and eliminate the released OSM-3CA, a process that depends on the engulfment receptor CED-1. Through genetic suppressor screens, we identified intragenic mutations in the OSM-3CA motor domain and mutations inhibiting the ciliary kinase DYF-5, both of which restored normal cilia in OSM-3CA-expressing animals. We showed that conformational changes in OSM-3CA prevent its entry into cilia, and OSM-3CA disposal requires its hyperactivity. Finally, we provide evidence that neurons also dispose of hyperactive kinesin-1 resulting from a clinic variant associated with amyotrophic lateral sclerosis, suggesting a widespread mechanism for regulating hyperactive kinesins.

Xie Q et al. (2005) J Biol Chem "Molecular cloning and characterization of a human AIF-like gene with ability to induce apoptosis."

Bonanno JA, Xie Q, Zheng J, Zhang Y, Lin T

[J Biol Chem, 2005]

In this study, we cloned and characterized a human gene homologous to the apoptosis-inducing factor (AIF), which is named AIF-like (AIFL). Human AIFL has 598 amino acids, with a characteristic Rieske domain and a pyridine nucleotide-disulfide oxidoreductase domain (Pyr_redox). AIFL shares 35% homology with AIF, mainly in the Pyr_redox domain. Reverse transcriptase-PCR analysis showed the expression of AIFL mRNA in all tissues tested, i.e. brain, colon, heart, kidney, liver, lung, muscle, ovary, pancreas, placenta, small intestine, and testis. We developed antibodies against human AIFL using fusion proteins as antigens. The antibodies specifically recognized the antigen and heterologously expressed AIFL proteins. The expression of AIFL proteins in human tissues was also ubiquitous, demonstrated by immunohistochemistry in tissue array slides. Subcellular fractionation and immunofluorescence staining studies revealed that AIFL is predominantly localized to the mitochondria. Similar to AIF, overexpression of AIFL induced apoptosis, as shown by increased cytoplasmic nucleosomes and subdiploid cell populations in AIFL-transfected cells. The segment 1-190 containing the Rieske domain induced apoptosis, whereas the segment containing the Pyr_redox domain did not contribute to the pro-apoptotic function. The mitochondrial membrane potential of cells transfected with AIFL was significantly more depolarized than that of the control. AIFL transfection-induced cytochrome c release and cleavage of caspase 3. Furthermore, the pan-caspase inhibitor Z-VAD-fmk inhibited AIFL induced apoptosis. In summary, AIFL induces apoptosis in a caspase-dependent manner when heterologously expressed.

Xie G et al. (2024) J Mol Histol "SPP-5 affects larval arrest via insulin signaling pathway in Caenorhabditis elegans."

Xie G, Shao Z

[J Mol Histol, 2024]

Diapause is an endocrine-mediated metabolic and growth arrest state in response to unfavorable external environments. The nematode Caenorhabditis elegans can enter diapause/arrest during embryonic, larval, or adult stages when subjected to detrimental external environments.&#xa0;Larval stage 1 (L1) arrest happens when animals hatch without food. Previous work has shown that the insulin pathway plays a prominent role in regulating L1 arrest. However, the downstream signal molecular mechanisms and biomarkers are still missing. In this study, we showed that SaPosin-like Protein family member SPP-5 is significantly upregulated during L1 arrest, suggesting that it could act as an L1 arrest biomarker. Using RNA interference we demonstrated that spp-5&#xa0; knockdown accelerated larval development, while the overexpression resulted in L1 arrest. Consistently,&#xa0;SPP-5 level was significantly up-regulated in the L1 arrest daf-2(e1370) mutants, and spp-5(RNAi) suppressed the daf-2(e1370) induced L1 arrest. These results suggest that SPP-5 can serve as an L1 arrest biomarker and promote the arrest probably via the insulin signaling pathway.

Xie X et al. (2020) J Vis Exp "Quantification of Insoluble Protein Aggregation in Caenorhabditis elegans during Aging with a Novel Data-Independent Acquisition Workflow."

Andersen JK, Angeli S, Xie X, Schilling B, Bhaumik D, Sivapatham R, Chamoli M, Lithgow GJ

[J Vis Exp, 2020]

We and others have shown that the aging process results in a proteome-wide accumulation of insoluble proteins. Knocking down genes encoding the insoluble proteins over 40% of the time results in an extension of the lifespan in C. elegans, suggesting that many of these proteins are key determinants of the aging process. Isolation and quantitative identification of these insoluble proteins are crucial to understand key biological processes that occur during aging. Here, we present a modified and improved protocol that details how to extract and isolate the SDS-insoluble proteins (insolublome) from C. elegans more efficiently to streamline mass spectrometric workflows via a novel label-free quantitative proteomics analysis. This improved protocol utilizes a highly efficient sonicator for worm lysis that greatly increases efficiency for protein extraction and allows us to use significantly less starting material (approximately 3,000 worms) than in previous protocols (typically using at least 40,000 worms). Subsequent quantitative proteomic analysis of the insolublome was performed using data-dependent acquisition (DDA) for protein discovery and identification and data-independent acquisition (DIA) for comprehensive and more accurate protein quantification. Bioinformatic analysis of quantified proteins provides potential candidates that can be easily followed up with other molecular methods in C. elegans. With this workflow, we routinely identify more than 1000 proteins and quantify more than 500 proteins. This new protocol enables efficient compound screening with C. elegans. Here, we validated and applied this improved protocol to wild-type C. elegans N2-Bristol strain and confirmed that aged day-10 N2 worms showed greater accumulation of the insolublome than day-2 young worms.