Joon Yong Kim

Functional genomic screen for modulators of ciliogenesis and cilium length

Primary cilia are evolutionarily conserved cellular organelles that organize diverse signalling pathways. Defects in the formation or function of primary cilia are associated with a spectrum of human diseases and developmental abnormalities. Genetic screens in model organisms have discovered core machineries of cilium assembly and maintenance. However, regulatory molecules that coordinate the biogenesis of primary cilia with other cellular processes, including cytoskeletal organization, vesicle trafficking and cell–cell adhesion, remain to be identified. Here we report the results of a functional genomic screen using RNA interference (RNAi) to identify human genes involved in ciliogenesis control. The screen identified 36 positive and 13 negative ciliogenesis modulators, which include molecules involved in actin dynamics and vesicle trafficking. Further investigation demonstrated that blocking actin assembly facilitates ciliogenesis by stabilizing the pericentrosomal preciliary compartment (PPC), a previously uncharacterized compact vesiculotubular structure storing transmembrane proteins destined for cilia during the early phase of ciliogenesis. The PPC was labelled by recycling endosome markers. Moreover, knockdown of modulators that are involved in the endocytic recycling pathway affected the formation of the PPC as well as ciliogenesis. Our results uncover a critical regulatory step that couples actin dynamics and endocytic recycling with ciliogenesis, and also provides potential target molecules for future study.

CEP290 interacts with the centriolar satellite component PCM-1 and is required for Rab8 localization to the primary cilium

Joubert syndrome (JS) is a developmental brain disorder characterized by cerebellar vermis hypoplasia, abnormal eye movement, ataxia and mental retardation. Mutations in CEP290 mutations are responsible for the cerebello-oculo-renal subtype of JS that includes kidney cysts and retinal degeneration, two phenotypes commonly linked to ciliopathies. CEP290 mutations are also associated with Meckel-Gruber syndrome and Bardet-Biedl syndrome (BBS). Here we demonstrate that CEP290 interacts with a centriolar satellite protein PCM-1, which is implicated in BBS4 function. CEP290 binds to PCM-1 and localizes to centriolar satellites in a PCM-1- and microtubule-dependent manner. The depletion of CEP290 disrupts subcellular distribution and protein complex formation of PCM-1. In accord with PCM-1s role in microtubule organization, CEP290 knockdown causes the disorganization of the cytoplasmic microtubule network. Moreover, we show that both CEP290 and PCM-1 are required for ciliogenesis and are involved in the ciliary targeting of Rab8, a small GTPase shown to collaborate with BBS protein complex to promote ciliogenesis. Our results suggest that PCM-1 is a potential mediator that may link CEP290 with BBS proteins in common molecular pathways.

Defective Wnt-dependent cerebellar midline fusion in a mouse model of Joubert syndrome

The ciliopathy Joubert syndrome is marked by cerebellar vermis hypoplasia, a phenotype for which the pathogenic mechanism is unclear. To investigate Joubert syndrome pathogenesis, we have examined mice with mutated Ahi1, the first identified Joubert syndrome–associated gene. These mice show cerebellar hypoplasia with a vermis-midline fusion defect early in development. This defect is concomitant with expansion of the roof plate and is also evident in a mouse mutant for another Joubert syndrome–associated gene, Cep290. Furthermore, fetal magnetic resonance imaging (MRI) of human subjects with Joubert syndrome reveals a similar midline cleft, suggesting parallel pathogenic mechanisms. Previous evidence has suggested a role for Jouberin (Jbn), the protein encoded by Ahi1, in canonical Wnt signaling. Consistent with this, we found decreased Wnt reporter activity at the site of hemisphere fusion in the developing cerebellum of Ahi1-mutant mice. This decrease was accompanied by reduced proliferation at the site of fusion. Finally, treatment with lithium, a Wnt pathway agonist, partially rescued this phenotype. Our findings implicate a defect in Wnt signaling in the cerebellar midline phenotype seen in Joubert syndrome that can be overcome with Wnt stimulation.

Extended-release formulation of morphine for subcutaneous administration

Pain arising from cancer tends to be chronic and chemotherapy of cancer pain usually requires narcotics. Most injectable narcotics, however, have short half-lives (T1/2) and require either continuous infusion or repeated frequent injections which are both inconvenient and uncomfortable. An extended-release formulation of morphine sulfate (Depo/Morphine) in a lipid-based drug-delivery system was characterized and tested in an animal model. The encapsulation efficiency was 53%±4%, and the in vitro release T1/2 in human plasma at 37° C was 12.1±1.1 days. Following s.c. administration of Depo/Morphine, the total amount of morphine remaining at the s.c. injection site decreased monoexponentially with a T1/2 value of 2.59±0.16 days as compared with 0.46±0.04 h following the injection of unencapsulated morphine. The morphine concentration in plasma also decreased monoexponentially with a T1/2 value of 8.33±2.13 days as compared with 0.45±0.21 h for unencapsulated morphine. Cataleptic behavior was observed in mice injected with unencapsulated morphine but not in those given an identical dose of morphine in the form of Depo/Morphine. In conclusion, Depo/Morphine has potential as an extended-release formulation of morphine and may be useful in chemotherapy of cancer pain as well as in maintenance therapy of narcotic addicts.

Genomic Analysis of a Pathogenic Bacterium, Paeniclostridium sordellii CBA7122 Containing the Highest Number of rRNA Operons, Isolated from a Human Stool Sample

Citation: Kim JY, Kim YB, Song HS, Chung W-H, Lee C, Ahn SW, Lee SH, Jung MY, Kim T-W, Nam Y-D and Roh SW (2017) Genomic Analysis of a Pathogenic Bacterium, Paeniclostridium sordellii CBA7122 Containing the Highest Number of rRNA Operons, Isolated from a Human Stool Sample. Front. Pharmacol. 8:840. doi: 10.3389/fphar.2017.00840 Genomic Analysis of a Pathogenic Bacterium, Paeniclostridium sordellii CBA7122 Containing the Highest Number of rRNA Operons, Isolated from a Human Stool Sample

Halorubrum aethiopicum sp. nov., an extremely halophilic archaeon isolated from commercial rock salt

A novel extremely halophilic archaeon, designated SAH-A6T, was isolated from a sample of commercial rock salt in Ethiopia. Cells of SAH-A6T were aerobic and pleomorphic. The strain was able to grow at concentrations of 15-30u200a% (w/v) NaCl (optimum 20-25u200a%u2009NaCl), at pH 6.0-9.0 (optimum pH 7.0) and in a temperature range of 30-55u2009°C (optimum 37-45u2009°C). Mg2+ was not required for growth of SAH-A6T cells. On the basis of 16S rRNA gene sequence analysis, strain SAH-A6T was closely related to Halorubrum halodurans Cb34T (99.1u200a%), Halorubrum rubrum YC87T (98.9u200a%), Halorubrum aquaticum EN-2T (98.7u200a%), Halorubrum cibi JCM 15757T (98.4u200a%), Halorubrum luteum CGSA15T (97.3u200a%), Halorubrum lipolyticum 9-3T (97.1u200a%), Halorubrum tibetense 8W8T (97.1u200a%), Halorubrum kocurii JCM 1478T (97.1u200a%), Halorubrum halophilum B8T (97.0u200a%) and Halorubrum persicum C49T (97.0u200a%). Phylogenetic analysis based on the rpoB gene sequences showed that strain SAH-A6T was closely related to Hrr. halodurans Cb34T (99.7u200a%), Hrr. aquaticum JCM 14031T (99.3u200a%) and other members of the genus Halorubrum (<99.0u200a%). The DNA G+Cu2009content of the strain was 68.0u2009mol%. DNA-DNA hybridization between strain SAH-A6T and the most closely related members of the genus Halorubrum were below 55u200a%, suggesting that the new isolate constitutes a different genospecies. On the bases of chemotaxonomic, phenotypic and genotypic data, strain SAH-A6T (=KCCM 43215T=JCM 31519T) represents a novel species of the genus Halorubrum, for which the name Halorubrumaethiopicum sp. nov. is proposed.

Complete genome sequence of Clostridium perfringens CBA7123 isolated from a faecal sample from Korea

BackgroundClostridium perfringens is an opportunistic human pathogen that causes necrotic enteritis, mild diarrhea, clostridial myonecrosis or gas gangrene, sepsis, etc. In this study, we aim to determine the pathogenesis of this bacterium at the genomic level. The genome of strain CBA7123 was sequenced, and a comparative genomic analysis between strain CBA7123 and four other related C. perfringens strains was performed.ResultsThe genome of strain CBA7123 consisted of one circular chromosome and one plasmid that were 3,088,370 and 46,640xa0bp long with 28.5 and 27.1xa0mol% G+C content, respectively. The genomic DNA was predicted to contain 2798 open reading frames (ORFs), 10 rRNA genes, and 94 tRNA genes. The genomic comparison analysis between the five strains revealed the distinctive virulence properties of strain CBA7123 by highlighting certain strain-specific genes.ConclusionsIn this study, the C. perfringens CBA7123 genome was sequenced and compared with other C. perfringens genomes. Among the various genes sequenced, the detection of antimicrobial resistance genes and those encoding various virulence factors may extend the understanding of the pathogenesis of C. perfringens strains.

Salicibibacter kimchii gen. nov., sp. nov., a moderately halophilic and alkalitolerant bacterium in the family Bacillaceae, isolated from kimchi

A moderately halophilic and alkalitolerant bacterial strain NKC1-1T was isolated from commercial kimchi in Korea. Strain NKC1-1T was Gram-stain-positive, aerobic, rod-shaped, non-motile, and contained diaminopimelic acid-type murein. Cell growth was observed in a medium containing 0–25% (w/v) NaCl (optimal at 10% [w/v]), at 20–40°C (optimal at 37°C) and pH 6.5–10.0 (optimal at pH 9.0). The major isoprenoid quinone of the isolate was menaquinone-7, and the major polar lipids were phosphatidylglycerol and unidentified phospholipids. Cell membrane of the strain contained iso-C17:0 and anteiso-C15:0 as the major fatty acids. Its DNA G + C content was 45.2 mol%. Phylogenetic analysis indicated the strain to be most closely related to Geomicrobium halophilum with 92.7–92.9% 16S rRNA gene sequence similarity. Based on polyphasic taxonomic evaluation with phenotypic, phylogenetic, and chemotaxonomic analyses, the strain represents a novel species in a new genus, for which the name Salicibibacter kimchii gen. nov., sp. nov. is proposed (= CECT 9537T; KCCM 43276T).

Role of jeotgal, a Korean traditional fermented fish sauce, in microbial dynamics and metabolite profiles during kimchi fermentation

We investigated the effects of jeotgal (fermented fish sauce) on kimchi fermentation, with or without saeu-jeot and myeolchi-jeot. Bacterial community analysis showed that Leuconostoc, Weissella, Lactobacillus, and Tetragenococcus were the dominant genera; however, their succession depended on the presence of jeotgal. Leuconostoc gasicomitatum was the dominant species in kimchi without jeotgal, whereas Weissella koreensis and Lactobacillus sakei were the dominant species in kimchi with myeolchi-jeot and saeu-jeot, respectively. Metabolite analysis, using 1H NMR, showed that the amounts of amino acids and gamma-aminobutyric acid (GABA) were higher in kimchi with jeotgal. Increases in acetate, lactate, and mannitol contents depended on fructose consumption and were more rapid in kimchi with jeotgal. Moreover, the consumption of various amino acids affected the increase in kimchi LAB. Thus, the role of jeotgal in kimchi fermentation was related to enhancement of taste, the amino acid source, and the increases in levels of functional metabolites.

Complete genome sequence of a commensal bacterium, Hafnia alvei CBA7124, isolated from human feces

BackgroundMembers of the genus Hafnia have been isolated from the feces of mammals, birds, reptiles, and fish, as well as from soil, water, sewage, and foods. Hafnia alvei is an opportunistic pathogen that has been implicated in intestinal and extraintestinal infections in humans. However, its pathogenicity is still unclear. In this study, we isolated H. alvei from human feces and performed sequencing as well as comparative genomic analysis to better understand its pathogenicity.ResultsThe genome of H. alvei CBA7124 comprised a single circular chromosome with 4,585,298xa0bp and a GC content of 48.8%. The genome contained 25 rRNA genes (9 5S rRNA genes, 8 16S rRNA genes, and 8 23S rRNA genes), 88 tRNA genes, and 4043 protein-coding genes. Using comparative genomic analysis, the genome of this strain was found to have 72 strain-specific singletons. The genome also contained genes for antibiotic and antimicrobial resistance, as well as toxin–antitoxin systems.ConclusionsWe revealed the complete genome sequence of the opportunistic gut pathogen, H. alvei CBA7124. We also performed comparative genomic analysis of the sequences in the genome of H. alvei CBA7124, and found that it contained strain-specific singletons, antibiotic resistance genes, and toxin–antitoxin systems. These results could improve our understanding of the pathogenicity and the mechanism behind the antibiotic resistance of H. alvei strains.