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By searching the mouse EST database, we identified a novel mouse cytosolic sulfotransferase (SULT) cDNA (RIKEN cDNA 2410078J06). Sequence analysis revealed that this new SULT belongs to the cytosolic SULT6 gene family. The recombinant form of this newly identified SULT, designated SULT6B1, was expressed using the pGEX-4T-1 glutathione S-transferase fusion system and purified from transformed BL21 Escherichia coli cells. Purified mouse SULT6B1 exhibited sulfonating activity toward thyroxine and bithionol among a variety of endogenous and xenobiotic compounds tested as substrates. pH optimum of purified mouse SULT6B1 was determined to be 8.0. Tissue-specific expression of mouse and human SULT6B1 was examined by RT-PCR. While human SULT6B1 was specifically expressed in kidney and testis, mouse SULT6B1 was detected in brain, heart, kidney, thymus, lung, liver and testis. Further studies are needed in order to clarify the role of SULT6B1 in the metabolism of thyroxine and possibly some xenobiotics in mouse.

EnteropathogenicEscherichia coli(EPEC) and enterohemorrhagicE. coli(EHEC) inject effector proteins into host cells via a type III secretion system encoded by the locus of enterocyte effacement (LEE). One of these effectors is Cif, encoded outside the LEE by a lambdoid prophage. In this study, we demonstrated that the Cif-encoding prophage of EPEC strain E22 is inducible and produces infectious phage particles. We investigated the distribution and functional expression of Cif in 5,049E. colistrains of human, animal, and environmental origins. A total of 115E. coliisolates from diverse origins and geographic locations carriedcif. The presence ofcifwas tightly associated with the LEE, since all thecif-positive isolates were positive for the LEE. These results suggested that the Cif-encoding prophages have been widely disseminated within the natural population ofE. colibut positively selected within the population of LEE-positive strains. Nonetheless, 66% ofcif-positiveE. colistrains did not induce a typical Cif-related phenotype in eukaryotic cells due to frameshift mutations or insertion of an IS element in thecifgene. The passenger region of the prophages carryingcifwas highly variable and showed various combinations of IS elements and genes coding for other effectors such asnleB,nleC,nleH,nleG,espJ, andnleA/espI(some of which were also truncated). This diversity and the presence of nonfunctional effectors should be taken into account to assess EPEC and EHEC pathogenicity and tropism.

At least three extracellular laminaran hydrolases which hydrolyzed laminaran (beta-1,3:1,6-glucan) from Eisenia bicyclis were secreted in wheat bran solid medium by Trichoderma viride U-1. These three enzymes, lam AI, AII, and B, were purified to electrophoretic homogeneity. Their molecular masses were estimated to be 70.1, 70.4, and 45.0 kDa for lam AI, AII, and B, respectively, by SDS-PAGE. Whereas both lam AI and AII could hydrolyze laminarin from Laminaria digitata, lam AII showed higher activity against Laminaria laminarin rather than Eisenia laminaran. On the other hand, lam B preferentially hydrolyzed pustulan, a beta-1,6-glucan. Laminarioligosaccharide was hydrolyzed by lam AI and AII but not B, whereas gentiooligosaccharide was hydrolyzed by only lam B. It showed that lam AI and AII were specific for beta-1,3-linkages, but lam B was specific for beta-1,6-linkages. These results indicated that T. viride U-1 has a multiple glucanolytic enzyme system.

RAPD and RFLP analyses were carried out to determine the origin of Livistona chinensis R. Br. var. subglobosa Becc. from Iriomotejima, Ishigakijima, Okinawa, Yakushima, Tanegashima, Cape Sata, Cape Toi, Tsukishima, and Aoshima, Japan. RAPD data obtained using 5 random primers were clustered using UPGMA or the neighbor-joining method. Each population was classified into three clusters based on the phylogenetic tree. L. chinensis plants from Yakushima, Tanegashima and Cape Sata belonged to the isologous cluster, and those from Ishigakijima and Okinawa are contained in a different cluster. L. chinensis from Okinawa and Ishigakijima could be differentiated by being the oldest based on the genetic distance. The area that covers Ishigakijima and Okinawa is thought to be the origin of L. chinensis. L. chinensis plants from Iriomotejima were contained in the same cluster as those from Aoshima. The phylogenetic trees constructed by both RAPD and RFLP analyses indicate the possibility that seeds or green woods of L. chinensis were dispersed by tidal current from the south field around Irlomotejima, and they were washed to Aoshima and established gradually. Therefore, we support the drifting-ashore-naturalized-plant hypothesis on the origin of L. chinensis in Aoshima.

By searching the GenBank database, we recently identified a novel mouse cytosolic sulfotransferase (SULT) cDNA (IMAGE Clone ID 679629) and a novel mouse SULT gene (LOC 215895). Sequence analysis revealed that both mouse SULTs belong to the cytosolic SULT3 gene family. The recombinant form of these two newly identified SULTs, designated SULT3A1 and SULT3A2, were expressed using the pGEX-4T-1 glutathione S-transferase fusion system and purified from transformed BL21 Escherichia coli cells. Both purified SULT3A1 and SULT3A2 exhibited strong amine N-sulfonating activities toward 1-naphthylamine among a variety of endogenous and xenobiotic compounds tested as substrates. Kinetic constants of the sulfation of 1-naphthylamine and 1-naphthol by these two enzymes were determined. Collectively, these results imply that these two amine-sulfonating SULT3s may play essential roles in the metabolism and detoxification of aromatic amine compounds in the body.

The gene lamAI, which encodes a novel laminarinase AI of Trichoderma viride U-1, was cloned using RT-PCR in conjunction with the rapid amplification of cDNA ends (RACE) technique. The open reading frame consisted of 2,277 bp encoding a protein of 759 amino acid residues, including a 32-residue signal prepropeptide. The protein showed 91% sequence similarity to the putative Trichoderma virens beta-1,3-glucanase BGN1, but no significant similarity to fungal beta-1,6-glucanases or beta-1,3-glucanases from other organisms. On 40 h incubation with a solo carbon source, northern analysis revealed that the gene was induced by 0.5% laminaran from Eisenia bicyclis but was not by the same concentration of glucose. The lamAI cDNA was functionally expressed in the methylotrophic yeast Pichia pastoris, resulting in a recombinant enzyme with as high activity against laminaran as native LAMAI. Based on these data, the probable existence of endo-beta-1,3:1,6-glucan hydrolases as a subclass of endo-beta-1,3-glucanases in some mycoparasitic fungi is suggested.

RAPD analysis was performed to discuss the origin of Livistona chinensis var. subglobosa using samples from eight localities, Iriomotejima, Ishigakijima, Okinawa, Yakushima, Tanegashima, Cape Sata, Tsukishima, and Aoshima, in Japan. Random amplified polymorphic DNA (RAPD) markers were obtained using five random primers and analyzed by the unweighted pair group method arithmetic (UPGMA). Data from Iriomotejima clustered with data from Aoshima, suggesting the possibility that seeds or green woods were carried by the tidal current from the southern fields of Iriomotejima to Aoshima.