PDLIM7 Synergizes With PDLIM2 and p62/Sqstm1 to Inhibit Inflammatory Signaling by Promoting Degradation of the p65 Subunit of NF-κB.

Activation of NF-κB transcription factors is critical for innate immune cells to induce inflammation and fight against microbial pathogens. On the other hand, the excessive and prolonged activation of NF-κB causes massive inflammatory damage to the host, suggesting that regulatory mechanisms to promptly terminate NF-κB activation are important to prevent immunopathology. We have previously reported that PDLIM2, a PDZ-LIM domain-containing protein, is a nuclear ubiquitin E3 ligase that targets the p65 subunit of NF-κB for degradation, thereby suppressing NF-κB activation. Here we show that PDLIM7, another member of LIM protein family, is also a ubiquitin E3 ligase that inhibits NF-κB-mediated inflammatory responses. PDLIM7 directly polyubiquitinates p65 and promotes its proteasomal degradation. Moreover, PDLIM7 heterodimerizes with PDLIM2 to promote synergistic PDLIM2-mediated degradation of p65. Mechanistically, PDLIM7 promotes K63-linked ubiquitination of PDLIM2 and then the proteasome/autophagosome cargo protein p62/Sqstm1 binds to both polyubiquitinated PDLIM2 and the proteasome, thereby facilitating the delivery of the NF-κB-PDLIM2 complex to the proteasome and subsequent p65 degradation. Consistently, double knockdown of PDLIM7 and either PDLIM2 or p62/Sqstm1 results in augmented proinflammatory cytokine production compared to control cells or single knockdown cells. These data delineate a new role for PDLIM7 and p62/Sqstm1 in the regulation of NF-κB signaling by bridging a ubiquitin E3 ligase and the proteasome.

HSP70 mediates degradation of the p65 subunit of nuclear factor κB to inhibit inflammatory signaling.

The nuclear PDZ-LIM domain protein PDLIM2 acts as a ubiquitin E3 ligase that targets the p65 subunit of the transcription factor nuclear factor κB (NF-κB) for degradation, thus preventing excessive inflammatory responses. We found that the chaperone protein HSP70 (heat shock protein of 70 kD) was required for the PDLIM2-mediated degradation of p65 and suppression of NF-κB signaling in lipopolysaccharide (LPS)-treated dendritic cells. In response to LPS, HSP70 translocated to the nucleus where it associated with PDLIM2 and the proteasome-associated protein BAG-1 (BCL2-associated athanogene 1) and promoted the transport of the NF-κB-PDLIM2 complex to the proteasome, thereby facilitating the degradation of p65. Consistent with these data, mouse dendritic cells deficient in either HSP70 or BAG-1 had more nuclear p65 and produced more proinflammatory cytokines than did wild-type dendritic cells. Furthermore, HSP70-deficient mice had more sustained inflammatory responses to bacterial infection than did wild-type mice. These data suggest that in addition to acting as a chaperone during protein folding, HSP70 plays a role in inhibiting proinflammatory NF-κB signaling by acting as a bridge between a ubiquitin E3 ligase and the proteasome.

Lectin-microarray technique for glycomic profiling of fungal cell surfaces.

Lectin microarrays are rows of lectins with different carbohydrate-binding specificities spotted on surfaces of glass slides. Lectin microarray technique enables glycomic analyses of carbohydrate composition of fungal cell walls. We will describe an application of the technique in analyzing cell surface glycome of yeast-form fungal cells in the living state. The analysis reveals genus- and species-dependent complex cell surface carbohydrate structures of fungi, and enabled us, therefore, to suggest that cell walls of yeast cells, which have been considered to have relatively simple structures, actually have a more complex structure containing galactose and fucose. This shows that the technique can be used to find new insights into the study of phylogenetic relations and into the classification of cells in the fungal kingdom based on cell wall glycome.

Human ZG16p recognizes pathogenic fungi through non-self polyvalent mannose in the digestive system.

Human zymogen granule protein 16 (ZG16p) contains a Jacalin-like lectin domain, although its glycan-binding properties are not fully understood. Here, we screened the glycan-binding specificity of ZG16p by recently developed glycoconjugate microarray. ZG16p appeared to exhibit selective binding to α- and ß-linked mannose-polyacrylamide-biotin probes. In more quantitative analysis using frontal affinity chromatography, dissociation constants to two types of polyvalent mannose, i.e. high-density mannose and yeast mannan, were determined to be 1.3 and 1.7 µM, respectively. Mutation of the evolutionarily conserved amino acid Asp151, which is involved in sugar binding among the Jacalin-related lectins (JRLs), abolished binding activity to mannose. By immunohistochemical staining, ZG16p was specifically detected in mucus-secreting cells of the digestive system such as serosanguineous acinar cells of the parotid gland, acinar cells of the pancreas and goblet cells of the intestine. Finally, we showed that ZG16p recognizes pathogenic Candida and Malassezia species in a polyvalent mannose-dependent manner. We propose that ZG16p is a novel member of mannose-specific JRLs, which recognizes pathogenic fungi through non-self polyvalent mannose in the digestive system.

Plasmacytoid dendritic cells are crucial for the initiation of inflammation and T cell immunity in vivo.

Plasmacytoid dendritic cells (pDCs) are characterized as type I interferon-producing cells that engage endosomal toll-like receptors (TLRs) and exclusively express sialic acid binding Ig-like lectin (Siglec)-H. However, their role in vivo remains unclear. Here we report a critical role for pDCs in the regulation of inflammation and T cell immunity in vivo by using gene-targeted mice with a deficiency of Siglec-H and conditional ablation of pDCs. pDCs were required for inflammation triggered by a TLR ligand as well as by bacterial and viral infections. pDCs controlled homeostasis of effector and regulatory CD4(+) T cells. Upon antigenic stimulation and microbial infection, pDCs suppressed the induction of CD4(+) T cell responses and participated in the initiation of CD8(+) T cell responses. Furthermore, Siglec-H appeared to modulate the function of pDCs in vivo. Thus, our findings highlight previously unidentified roles of pDCs and the regulation of their function for the control of innate and adaptive immunity.

Yojironins E-I, prenylated acylphloroglucinols from Hypericum yojiroanum.

Five new prenylated acylphloroglucinols, yojironins E-Ι (1-5), were isolated from the whole plants of Hypericum yojiroanum. Their structures were elucidated by spectroscopic data. Yojironin E (1) exhibited antimicrobial activity against Aspergillus niger, Candida albicans, Cryptococcus neoformans, and Trichophyton mentagrophytes.

Nakijinamines C-E, new heteroaromatic alkaloids from the sponge Suberites species.

Three new heteroaromatic alkaloids, nakijinamines C-E (1-3), which are a hybrid of the aaptamine-type and bromoindole alkaloids possessing a taurine- or histidine-derived residue, have been isolated from an Okinawan marine sponge Suberites species. The structures of 1-3 were elucidated on the basis of spectroscopic data and chemical conversions. Nakijinamines C (1) and E (3) are the first natural products possessing a 1H-oxazolo[4',5':4,5]benzo[1,2,3-de][1,6]naphthyridine ring system.

Streptomyces coacervatus sp. nov., isolated from the intestinal tract of Armadillidium vulgare.

A Gram-staining-positive bacterium, designated AS-0823(T), which formed spiral spore chains on the aerial mycelium, was isolated from the intestinal tract of Armadillidium vulgare, a small terrestrial crustacean commonly found on the ground around houses in Japan. 16S rRNA gene sequence analysis showed that the isolate belonged to the genus Streptomyces and was most closely related to Streptomyces longisporus ISP 5166(T) (98.6 % 16S rRNA gene sequence similarity), Streptomyces curacoi NBRC 12761(T) (98.4 %) and Streptomyces griseoruber NBRC 12873(T) (98.4 %). The affiliation of strain AS-0823(T) to the genus Streptomyces was supported by chemotaxonomic data: iso-C(16 : 0), anteiso-C(15 : 0), C(16 : 0), iso-C(15 : 0) and anteiso-C(17 : 0) as the major cellular fatty acids, ll-diaminopimelic acid as the characteristic diamino acid in the peptidoglycan and the absence of mycolic acids. DNA-DNA hybridization and physiological and biochemical analysis supported the differentiation of strain AS-0823(T) from S. longisporus JCM 4395(T). Therefore, strain AS-0823(T) represents a novel species, for which the name Streptomyces coacervatus sp. nov. is proposed. The type strain is AS-0823(T) ( = IFM 11055(T)  = DSM 41983(T)  = JCM 17138(T)).

Petiolins J-M, prenylated acylphloroglucinols from Hypericum pseudopetiolatum var. kiusianum.

Four new prenylated acylphloroglucinols, petiolins J-M (1-4), were isolated from aerial parts of Hypericum pseudopetiolatum var. kiusianum, and the structures were elucidated by spectroscopic data and a single-crystal X-ray diffraction analysis. Petiolin J (1) exhibited antimicrobial activity.

Ceratinadins A-C, new bromotyrosine alkaloids from an Okinawan marine sponge Pseudoceratina sp.

Three new bromotyrosine alkaloids, ceratinadins A-C (1-3), were isolated from an Okinawan marine sponge Pseudoceratina sp. and the structures of 1-3 were elucidated on the basis of spectroscopic data. Ceratinadin A (1) was a novel bromotyrosine alkaloid possessing an N-imidazolyl-quinolinone moiety. Ceratinadins A (1) and B (2) showed antifungal activity.