The synaptonemal complex is assembled by a polySUMOylation-driven feedback mechanism in yeast

During meiotic prophase I, proteinaceous structures called synaptonemal complexes (SCs) connect homologous chromosomes along their lengths via polymeric arrays of transverse filaments (TFs). Thus, control of TF polymerization is central to SC formation. Using budding yeast, we show that efficiency of TF polymerization closely correlates with the extent of SUMO conjugation to Ecm11, a component of SCs. HyperSUMOylation of Ecm11 leads to highly aggregative TFs, causing frequent assembly of extrachromosomal structures. In contrast, hypoSUMOylation leads to discontinuous, fragmented SCs, indicative of defective TF polymerization. We further show that the N terminus of the yeast TF, Zip1, serves as an activator for Ecm11 SUMOylation. Coexpression of the Zip1 N terminus and Gmc2, a binding partner of Ecm11, is sufficient to induce robust polySUMOylation of Ecm11 in nonmeiotic cells. Because TF assembly is mediated through N-terminal head-to-head associations, our results suggest that mutual activation between TF assembly and Ecm11 polySUMOylation acts as a positive feedback loop that underpins SC assembly.     

Protection against metastasis by immunization with an allogeneic lymphocyte antigen

Q5 antigens are expressed on the surface of various experimental murine tumor cells. They share partially common antigenicity with Qa-2 alloantigens expressed on normal lymphocytes. For that reason we tested the immunoprotection by anti-Qa-2 immunization of mice against a Q5+ tumor. Nerve fibrosarcoma (NSFA) tumor, which specifically develops metastasis in the lung, has been reported to be poorly immunogenic. However, expression of the Q5 antigen was evident on the surface of NFSA cells. After immunizing (C3H/He x B6.K1)F1 (Qa-2-) mice with B6 (Qa-2+) lymphocytes, the protection against the proliferation of the semi-syngeneic NFSA tumor was examined First, immunization of normal mice induced resistance to NFSA cell transplants. Second, when the tumor cells were transplanted to the hind foot of a mouse and the resulting tumor was removed by amputating the leg, the mice were protected against the development of lung metastasis after immunization by intraperitoneal inoculation of B6 cells 3 days after tumor removal. Immunization with attenuated NFSA cells in this system failed to protect the mice from lung metastasis. On the other hand, inoculation of the mice with B6 cells without removal of the original tumor on the foot showed little effect on the progression of the tumor. Thus, cytotoxic T lymphocytes (CTL), which seemed to be present in an inactive form in the mice from which the tumor had not been removed, were induced in the mice after the removal of the major tumor followed by immunization with B6 lymphocytes. The induction of CTL by the immunization was suppressed in mice bearing large tumors. Cells stimulated by the tumor antigen seemed to be involved in the suppression. It was also shown that the Q5 antigen is the direct recognition target of the CTL since the activity of Q5-specific CTL clones in lysing tumor cells was inhibited by a monoclonal antibody specific for the Q5 antigen. In contrast to immunization with attenuated tumor cells, our novel allogeneic lymphocyte immunization procedure offers high CTL activation, by-passing the induction of T cell unresponsiveness.     

Development and characterization of 21 polymorphic microsatellite loci in the aphidophagous gall midge, <Emphasis Type="Italic">Aphidoletes aphidimyza</Emphasis> (Diptera: Cecidomyiidae)

We have developed and characterized 21 microsatellite markers in the aphidophagous gall midge Aphidoletes aphidimyza (Rondani) (Diptera: Cecidomyiidae). All 21 loci tested were polymorphic: the number of alleles ranged from 2 to 17. Allelic richness and observed heterozygosities were higher in females than in males. Several loci had no heterozygosity in males, suggesting that the loci were located on sex chromosomes or E-chromosomes, common to cecidomyiids. The high polymorphism detected in this study suggests the markers will be of value in analyzing genetic structure of field populations.     

Hydrophobic Components in Delipidated Granule of Egg Yolk

Dimethyl sulfide (DMS) is volatile compound important as one of the characteristic flavor compounds of marine food products. The precursor of DMS in marine products is dimethyl-β- propiothetin (DMPT), which is abundunt in green algae. DMPT was effectively extracted by the use of hydrophilic solvents from dried hitoegusa (Monostroma nitidum), a green alga. At around pH 4 and at over pH 9, the extracted DMPT was rapidly degraded to DMS; at around pH 7.5, this degradation was much slower. The DMS obtained volatilized immediately from aqueous solution. However, when the DMPT was formed into a powder with dextrin and heated to release the DMS, 40 — 60% of the DMS remained in the powder. The amount of DMS remaining was 80 % when cyclodextrin was used to form the powder.     

Colocalization of apolipoprotein AI in various kinds of systemic amyloidosis.

Apolipoprotein AI (apoAI), a major component of high-density lipoproteins, is one of the major amyloid fibril proteins and a minor constituent of the senile plaques observed in Alzheimer's disease. We examined colocalization of apoAI in various kinds of systemic amyloidosis in this study. Forty-three of 48 formalin-fixed paraffin-embedded heart specimens with various forms of systemic amyloidosis reacted immunohistochemically with anti-human apoAI antibody. ApoAI was also detected in water-extracted amyloid material by immunoblotting. In addition, we observed colocalization of apoAI and murine amyloid A (AA) amyloidosis in human apoAI transgenic mice. This is the first report of colocalization of apoAI with amyloid deposits in various forms of human systemic amyloidosis and murine AA amyloidosis in human apoAI transgenic mice. ApoAI may not always be a major component of amyloid fibrils, even when it is present in systemic amyloid deposits.     

A chondroitin synthase-1 (ChSy-1) missense mutation in a patient with neuropathy impairs the elongation of chondroitin sulfate chains initiated by chondroitin N-acetylgalactosaminyltransferase-1

Background

Previously, we identified two missense mutations in the chondroitin N-acetylgalactosaminyltransferase-1 gene in patients with neuropathy. These mutations are associated with a profound decrease in chondroitin N-acetylgalactosaminyltransferase-1 enzyme activity. Here, we describe a patient with neuropathy who is heterozygous for a chondroitin synthase-1 mutation. Chondroitin synthase-1 has two glycosyltransferase activities: it acts as a GlcUA and a GalNAc transferase and is responsible for adding repeated disaccharide units to growing chondroitin sulfate chains.

Methods

Recombinant wild-type chondroitin synthase-1 enzyme and the F362S mutant were expressed. These enzymes and cells expressing them were then characterized.

Results

The mutant chondroitin synthase-1 protein retained approximately 50% of each glycosyltransferase activity relative to the wild-type chondroitin synthase-1 protein. Furthermore, unlike chondroitin polymerase comprised of wild-type chondroitin synthase-1 protein, the non-reducing terminal 4-O-sulfation of GalNAc residues synthesized by chondroitin N-acetylgalactosaminyltransferase-1 did not facilitate the elongation of chondroitin sulfate chains when chondroitin polymerase that consists of the mutant chondroitin synthase-1 protein was used as the enzyme source.

Conclusions

The chondroitin synthase-1 F362S mutation in a patient with neuropathy resulted in a decrease in chondroitin polymerization activity and the mutant protein was defective in regulating the number of chondroitin sulfate chains via chondroitin N-acetylgalactosaminyltransferase-1. Thus, the progression of peripheral neuropathies may result from defects in these regulatory systems.

General significance

The elongation of chondroitin sulfate chains may be tightly regulated by the cooperative expression of chondroitin synthase-1 and chondroitin N-acetylgalactosaminyltransferase-1 in peripheral neurons and peripheral neuropathies may result from synthesis of abnormally truncated chondroitin sulfate chains.     

Unique features of the motility and structures in the flagellate polar region of Campylobacter jejuni and other species: an electron microscopic study

Similarly to Helicobacter pylori but unlike Vibrio cholerae O1/O139, Campylobacter jejuni is non‐motile at 20°C but highly motile at ≥37°C. The bacterium C. jejuni has one of the highest swimming speeds reported (>100 μm/s), especially at 42°C. Straight and spiral bacterial shapes share the same motility. C. jejuni has a unique structure in the flagellate polar region, which is characterized by a cup‐like structure (beneath the inner membrane), a funnel shape (opening onto the polar surface) and less dense space (cytoplasm). Other Campylobacter species (coli, fetus, and lari) have similar motility and flagellate polar structures, albeit with slight differences. This is especially true for Campylobacter fetus, which has a flagellum only at one pole and a cup‐like structure composed of two membranes.     

Overexpression of mutant cell division cycle 25 homolog B (CDC25B) enhances the efficiency of selection in Chinese hamster ovary cells

The effects of mutant cell division cycle 25 homolog B (CDC25B) overexpression on the generation of cells producing a monoclonal antibody were investigated in Chinese hamster ovary (CHO) cells. Mutant CDC25B (m-CDC25B) expression plasmids were transfected into CHO DG44-derived cells producing a monoclonal antibody, and the frequency of highly producing cells was assessed following gene amplification in the presence of 250 nM methotrexate. Most of the clones obtained from the m-CDC25B-overexpressing cells had higher antibody titers than did mock-transfected control cells. This arose from either higher transgene copy numbers or higher mRNA expression levels for the antibody. However, the high mRNA expression levels were not always accompanied by increases in transgene copy numbers. Our results suggest that cells producing high levels of a monoclonal antibody can be selected efficiently using m-CDC25B overexpression.     

Possible involvement of Hcn1 ion channel in learning and memory dysfunction in SAMP8 mice

The senescence-accelerated mouse prone 8 (SAMP8) strain exhibits age-related learning and memory deficits (LMD) at 2 months of age. Combined linkage analysis of 264 F2 intercross SAMP8 × JF1 mice and RNA-seq analysis identified Hcn1 gene out of 29 genes in the LMD region on chromosome 13. Hcn1 in SAMP8 strain showed 15 times less polyglutamine repetition compared to Japanese fancy mouse 1 (JF1). Whole cell patch clamp analysis showed that Hcn1 ion conductivity was significantly lower in SAMP8 compared to that of JF1, which may be associated with learning and memory deficiency.     

Sulfation of keratan sulfate proteoglycan reduces radiation-induced apoptosis in human Burkitt’s lymphoma cell lines

This study focuses on clarifying the contribution of sulfation to radiation-induced apoptosis in human Burkitt’s lymphoma cell lines, using 3′-phosphoadenosine 5′-phosphosulfate transporters (PAPSTs). Overexpression of PAPST1 or PAPST2 reduced radiation-induced apoptosis in Namalwa cells, whereas the repression of PAPST1 expression enhanced apoptosis. Inhibition of PAPST slightly decreased keratan sulfate (KS) expression, so that depletion of KS significantly increased radiation-induced apoptosis. In addition, the repression of all three N-acetylglucosamine-6-O-sulfotransferases (CHST2, CHST6, and CHST7) increased apoptosis. In contrast, PAPST1 expression promoted the phosphorylation of p38 MAPK and Akt in irradiated Namalwa cells. These findings suggest that 6-O-sulfation of GlcNAc residues in KS reduces radiation-induced apoptosis of human Burkitt’s lymphoma cells.