Preparation of elastic silk sericin hydrogel

This paper reports a preparation method for silk sericin hydrogel using the Sericin-hope silkworm, whose cocoons consist almost exclusively of sericin. Sericin solution, prepared from Sericin-hope cocoons, contains intact sericin and forms elastic hydrogels with the addition of ethanol. The sericin hydrogel can be prepared without crosslinking by chemicals or irradiation and might be usable as a naturally occurring biomaterial.     

CLASP1 and CLASP2 bind to EB1 and regulate microtubule plus-end dynamics at the cell cortex

CLIP-associating protein (CLASP) 1 and CLASP2 are mammalian microtubule (MT) plus-end binding proteins, which associate with CLIP-170 and CLIP-115. Using RNA interference in HeLa cells, we show that the two CLASPs play redundant roles in regulating the density, length distribution and stability of interphase MTs. In HeLa cells, both CLASPs concentrate on the distal MT ends in a narrow region at the cell margin. CLASPs stabilize MTs by promoting pauses and restricting MT growth and shortening episodes to this peripheral cell region. We demonstrate that the middle part of CLASPs binds directly to EB1 and to MTs. Furthermore, we show that the association of CLASP2 with the cell cortex is MT independent and relies on its COOH-terminal domain. Both EB1- and cortex-binding domains of CLASP are required to promote MT stability. We propose that CLASPs can mediate interactions between MT plus ends and the cell cortex and act as local rescue factors, possibly through forming a complex with EB1 at MT tips.     

Identification of the pathogenic Aspergillus species by nested PCR using a mixture of specific primers to DNA topoisomerase II gene

For PCR-based identification of Aspergillus species, a common primer of the DNA topoisomerase II genes of Candida, Aspergillus and Penicillium, and species-specific primers of the genomic sequences of DNA topoisomerase II of A. fumigatus, A. niger, A. flavus (A. oryzae), A. nidulans and A. terreus were tested for their specificities in PCR amplifications. The method consisted of amplification of the genomic DNA topoisomerase II gene by a common primer set, followed by a second PCR with a primer mix consisting of 5 species-specific primer pairs for each Aspergillus species. By using the common primer pair, a DNA fragment of approximately 1,200 bp was amplified from the Aspergillus and Penicillium genomic DNAs. Using each species-specific primer pair, unique sizes of PCR products were amplified, all of which corresponded to a species of Aspergillus even in the presence of DNAs of several fungal species. The sensitivity of A. fumigatus to the nested PCR was found to be 100 fg of DNA in the reaction mixture. In the nested PCR obtained by using the primer mix (PsIV), the specific DNA fragment of A. fumigatus was amplified from clinical specimens. These results suggest that this nested PCR method is rapid, simple and available as a tool for identification of pathogenic Aspergillus to a species level.     

A new species of Megoura (Hemiptera,Aphididae) from Japan

A new species of the genus Megoura, M. lathyricola sp. n., was collected from Lathyrus japonicus subsp. japonicus (Leguminosae) in seashore areas of northern and southern Japan. This species is described and illustrated, and a revised key to the identification of the world species of Megoura is presented.     

Identification of a link between the SAMP repeats of adenomatous polyposis coli tumor suppressor and the Src homology 3 domain of DDEF

The adenomatous polyposis coli (APC) tumor suppressor protein is a multifunctional protein with a well characterized role in the Wnt signal transduction pathway and in cytoskeletal regulation. The SAMP repeats region of APC, an Axin-binding site, is known to be important for tumor suppression and for the developmental function of APC. We performed a yeast two-hybrid screening using the first SAMP motif-containing region of Xenopus APC as bait and obtained several SAMP binding candidates including DDEF2 (development and differentiation enhancing factor 2), which is an ADP-ribosylation factor (Arf) GTPase-activating protein (GAP (ArfGAP)) involved in the regulation of focal adhesions. In vitro and in cells the Src homology 3 (SH3) domain of DDEF2 and its close homolog, DDEF1, are associated with the SAMP motif of APC competitively with Axin1. Moreover, NMR chemical shift perturbation experiments revealed that the SAMP motif interacts at the same surface of the SH3 domain of DDEF as the known SH3 binding motif, PXXP. When fluorescent protein-tagged APC and DDEF are expressed in Xenopus A6 cells, co-localization at microtubule ends is observed. Overexpression and RNA interference experiments indicate that APC and DDEFs cooperatively regulate the distributions of microtubules and focal adhesions. Our findings reveal that the SAMP motif of APC specifically binds to the SH3 domains of DDEFs, providing new insights into the functions of APC in cell migration.     

Effects of a casein hydrolysate prepared from Aspergillus oryzae protease on adjuvant arthritis in rats

We evaluated the effects of a casein hydrolysate (CH) prepared from Aspergillus oryzae protease on rat adjuvant arthritis, a model of human rheumatoid arthritis. CH was administered orally once a day to the animals for 22 d after the adjuvant injection. CH suppressed swelling in the adjuvant-uninjected hind paws, and a higher dose of CH suppressed the increase in arthritic score and swelling of the adjuvant-injected hind paws. A histopathological examination revealed evidence that the higher dose of CH suppressed the articular changes in the rats. In addition, CH suppressed the production of nitric oxide and prostaglandin E(2) in the plasma of the rats. These results suggest that CH had a suppressive effect on adjuvant arthritis by inhibiting the acute and chronic inflammatory reactions.     

Identification of a single nuclear localization signal in the C-terminal domain of an Aspergillus DNA topoisomerase II

DNA topoisomerase II (topo II) is a major nuclear protein that plays an important role in DNA metabolism. We have isolated the gene for topo II ( TOP2) from the filamentous fungus Aspergillus terreus. The deduced amino acid sequence revealed that topo II consists of 1,587 amino acids and has a calculated molecular weight of 180 kDa; the protein expressed in Escherichia coli has an estimated molecular weight of 185 kDa. Expression of topo II polypeptides tagged with yellow fluorescent protein (YFP) in budding yeast suggests that the C-terminal region of the topo II is essential for transport of the fusion protein into the nucleus. The nuclear localization signal (NLS) sequence of topo II is a non-classical bipartite type containing two interdependent, positively charged clusters separated by 15 amino acids. Alanine scanning mutagenesis and deletion analyses showed further that a stretch of 23 amino acid residues (positions 1,234-1,256) is necessary for nuclear import. In addition, we confirmed, using co-immunoprecipitation and two-hybrid analysis, that this non-classical NLS interacts with importin alpha in budding yeast. These results suggest that the fungal topo II NLS is functional in yeast cells.