Purification and Properties of Tyrosinase Isozymes from the Gill of Lentinus edodes Fruiting Body

Six tyrosinase isozymes were purified from the browned gill of the fruiting body of Lentinus edodes by ammonium sulfate fractionation, DEAE-Sephacel and Q-Sepharose column chromatography, and partially denaturing SDS–PAGE. At the step of Q-Sepharose column chromatography, two active fractions (A and B) were obtained. Each fraction was separated to three further fractions, A1, A2, and A3, and B1, B2, and B3, respectively, by partially denaturing SDS–PAGE. All these isozymes consisted of two types of polypeptides: a polypeptide (Aα or Bα) and either β (Aβ or Bβ) or γ polypeptide (Aγ or Bγ). The α polypeptide contained the consensus amino acid sequence of the active site of known tyrosinases, which is considered to act as a catalytic subunit. From the results of peptide mapping and the amino acid composition, Aα and Bα polypeptides were considered to be different proteins. The kinetic properties of the purified tyrosinase isozymes differed greatly according to whether they contained β or γ polypeptide, indicating these polypeptides to be a possible regulatory subunit.     

Cloning of the Lentinula edodes B mating-type locus and identification of the genetic structure controlling B mating

During the life cycle of heterothallic tetrapolar Agaricomycetes such as Lentinula edodes (Berk.) Pegler, the mating type system, composed of unlinked A and B loci, plays a vital role in controlling sexual development and resulting formation of the fruit body. L. edodes is produced worldwide for consumption and medicinal purposes, and understanding its sexual development is therefore of great importance. A considerable amount of mating type factors has been indicated over the past decades but few genes have actually been identified, and no complete genetic structures of L. edodes B mating-type loci are available. In this study, we cloned the matB regions from two mating compatible L. edodes strains, 939P26 and 939P42. Four pheromone receptors were identified on each new matB region, together with three and four pheromone precursor genes in the respective strains. Gene polymorphism, phylogenetic analysis and distribution of pheromone receptors and pheromone precursors clearly indicate a bipartite matB locus, each sublocus containing a pheromone receptor and one or two pheromone precursors. Detailed sequence comparisons of genetic structures between the matB regions of strains 939P42, 939P26 and a previously reported strain SUP2 further supported this model and allowed identification of the B mating type subloci borders. Mating studies confirmed the control of B mating by the identified pheromone receptors and pheromones in L. edodes.     

Murine sex-limited protein (Slp) antigenic sites in human complement component C4

Human complement component C4 is encoded by two HLA-linked loci, A and B. In the mouse, the H-2 region contains structural genes for two serum proteins that react with antibodies to human C4, but one of these proteins (Slp) has no C4 hemolytic activity. Because the product of C4-A locus in man has low hemolytic activity, a previous report suggests it may be the homologue of murine Slp. We show here that Slp antigenic determinants are found in human C4. However, they are expressed in the products of both loci A and B, that is, C4A and C4B, since both proteins were specifically immunoprecipitated by the IgG fraction of alloantisera to mouse Slp. Therefore, Slp-associated structural features are preserved in evolution, although they do not seem to be relevant to the hemolytic properties of C4.     

Paternal phylogeography and genetic diversity of East Asian goats

This study was a first analysis of paternal genetic diversity for extensive Asian domestic goats using SRY gene sequences. Sequencing comparison of the SRY 3′‐untranslated region among 210 Asian goats revealed four haplotypes (Y1A, Y1B, Y2A and Y2B) derived from four variable sites including a novel substitution detected in this study. In Asian goats, the predominant haplotype was Y1A (62%) and second most common was Y2B (30%). Interestingly, the Y2B was a unique East Asian Y chromosomal variant, which differentiates eastern and western Eurasian goats. The SRY geographic distribution in Myanmar and Cambodia indicated predominant the haplotype Y1A in plains areas and a high frequency of Y2B in mountain areas. The results suggest recent genetic infiltration of modern breeds into South‐East Asian goats and an ancestral SRY Y2B haplotype in Asian native goats.     

Hordein Polymorphism in Ethiopian Barley

Using starch gel electrophoresis, polymorphism of hordein-encoding loci Hrd A, Hrd B, and Hrd Fwas studied in 147 accessions of local Ethiopian barley cultivars. Loci Hrd A, Hrd B, and Hrd Fwere shown to have 26, 36, and 4 alleles, respectively. The allele frequencies in the collection examined varied from 0.17 to 45.72%. For loci Hrd Aand Hrd B, families of blocks of hordein components were found. Based on the allele frequencies and their combinations at loci Hrd Aand Hrd Bas well as the numbers of families of component blocks of hordeins A and B, we identified genotypes that could be considered as the most ancient in Ethiopia. A catalog of hordein variant encoded by these loci was created. The list of hordein genetic formulas for the studied accessions is presented.     

Molecular markers linked to genes affecting plant height in wheat using a doubled-haploid population

 Plant height in wheat (Triticum aestivum L. em Thell) is known to be under polygenic control. Crosses involving genes Rht-B1 and Rht-D1, located on chromosomes 4BS and 4DS, respectively, have shown that these genes have major effects. Two RFLP loci were found to be linked to these two genes (Xfba1-4B with Rht-B1 and Xfba211-4D with Rht-D1) by genotyping a population of F₁-derived doubled-haploid lines [‘Courtot’ (Rht-B1b+Rht-D1b)בChinese Spring’]. Using a well-covered molecular marker map, we detected three additional regions and one interaction influencing plant height. These regions, located on chromosome arms 4BS (near the locus Xglk556-4B), 7AL (near the locus Xglk478-7A) and 7BL (near the locus XksuD2-7B) explained between 5% and 20% of the variability for this trait in this cross. The influence of 2 loci from chromosome 4B (Xfba1-4B and Xglk556-4B) suggests that there could be a duplication of Rht-B1 on this chromosome originating from Cv ‘Courtot’. Moreover, an interaction effect between loci from chromosome arms 1AS (near the locus Xfba393-1A) and 1BL (near the locus Xcdo1188-1B) was comparable to or even higher than those of the Rht-B1b and Rht-D1b alleles. A model including the main effects of the loci from chromosomes 4B and 4D (Xfba1-4B, Xglk556-4B and Xfba211-4D) and the interaction effect between Xfba393-1A and Xcdo1188-1B is proposed, which explains about 50% of the variation in plant height. The present results are discussed in relation to those obtained using nullisomic or substitution lines. Received: 13 June 1997 / Accepted: 13 October 1997     

UBE3A-mediated regulation of imprinted genes and epigenome-wide marks in human neurons

The dysregulation of genes in neurodevelopmental disorders that lead to social and cognitive phenotypes is a complex, multilayered process involving both genetics and epigenetics. Parent-of-origin effects of deletion and duplication of the 15q11-q13 locus leading to Angelman, Prader-Willi, and Dup15q syndromes are due to imprinted genes, including UBE3A, which is maternally expressed exclusively in neurons. UBE3A encodes a ubiquitin E3 ligase protein with multiple downstream targets, including RING1B, which in turn monoubiquitinates histone variant H2A.Z. To understand the impact of neuronal UBE3A levels on epigenome-wide marks of DNA methylation, histone variant H2A.Z positioning, active H3K4me3 promoter marks, and gene expression, we took a multi-layered genomics approach. We performed an siRNA knockdown of UBE3A in two human neuroblastoma cell lines, including parental SH-SY5Y and the SH(15M) model of Dup15q. Genes differentially methylated across cells with differing UBE3A levels were enriched for functions in gene regulation, DNA binding, and brain morphology. Importantly, we found that altering UBE3A levels had a profound epigenetic effect on the methylation levels of up to half of known imprinted genes. Genes with differential H2A.Z peaks in SH(15M) compared to SH-SY5Y were enriched for ubiquitin and protease functions and associated with autism, hypoactivity, and energy expenditure. Together, these results support a genome-wide epigenetic consequence of altered UBE3A levels in neurons and suggest that UBE3A regulates an imprinted gene network involving DNA methylation patterning and H2A.Z deposition.     

Differential expression of activin βA and βB genes in female allotriploid and diploid red crucian carp Carassius auratus red var

activin β_A and β_B from diploid and allotriploid crucian carp were cloned.The differential expression of activin β_A and β_B genes in female allotriploid and diploid red crucian carp Carassius auratus red var. were studied and found to be expressed in all the tested tissues; particularly, the expression of activin β_A and β_B was elevated in the ovaries of allotriploids and differential expression in pituitaries during the non-breeding season and the breeding season period. The immunohistochemistry indicated that the abnormal triploid ovaries were dominated by small oogonium-like cells with dense signals and that the elevated expression of activin β_A and β_B in the ovaries of allotriploids may be related to allotriploid sterility.     

Regions in theZ5 mating gene ofSchizophyllum commune involved in Y-Z binding and recognition

The Aα mating locus of the woodrotting fungusSchizophyllum commune encodes two multiallelic genes,Y andZ, which regulate the A-pathway of development. TheY alleles contain a homeobox, suggesting that the Y proteins may be DNA-binding regulatory proteins. During mating, development is induced when Y from one mating partner interacts with Z from the other mating partner; self combinations of Y and Z are inactive. Two-hybrid analyses indicate that nonself combinations of Y and Z form heteromultimers and self combinations do not. To understand Y-Z binding and self- nonself recognition further we used mutagenesis and chimeras to identify regions in one allele ofZ(Z5) that are involved in these processes. Here we report the results, which broadly define regions in Z5 that are essential for activity, Y-Z binding and Z5 allelic specificity.     

Genomic and Yeast Artificial Chromosome Long-Range Regular Article Linking Six Loci in 10q11.2 and Spanning the Multiple Endocrine Neoplasia Type 2A (MEN2A) Region

Multiple endocrine neoplasia types 2A and 2B (MEN 2A and MEN 2B) and familial medullary thyroid carcinoma (FMTC) are dominantly inherited cancers that have in common the clinical feature of medullary thyroid carcinoma (MTC). We have performed both genomic long-range restriction mapping and yeast artificial chromosome (YAC) contig assembly and restriction mapping to establish physical linkage, order, and distances between six loci in 10q11.2 near the genes responsible for these hereditary cancers. RET, D10S94, D10S182, and D10S102 have been mapped in genomic DNA. RET, D10S94, D10S182, D10F3853, and the 10q11.2 sequences detected by DNA marker DM124 are encompassed by a 1-Mb YAC contig. Six physically linked loci are within 1.4 Mb and have an order and orientation of 10cen, D10F38S3, DM124, RET, D10S94, D10S182, D10S102, 10qter. Mutations in the RET proto-oncogene have recently been demonstrated to be associated with MEN 2A and FMTC. RET is located within a genetically defined MEN2A candidate interval between D10S141 and D10S94; MEN2B has been mapped to a larger, overlapping region between D10S141 and a more distal locus, RBP3. Both our genomic physical map and our YAC contig span the entire MEN2A candidate region and overlap with that of MEN2B . These maps will facilitate the identification of genes that can be considered candidates for MEN2B and the identification of tumor-specific alterations important in sporadic MTC.     

担子菌交配型基因的克隆及功能研究进展

担子菌中已有２０多个交配型基因被克隆,Ａ、Ｂ位点的交配型基因具有明显不同的结构特征。Ａ位点的交配型基因编码两类含相似同源结构域的蛋白质,两种蛋白质形成异源二聚体,调节依赖Ａ因子的发育过程;而Ｂ位点的交配型基因编码信息素及其受体。交配型基因编码的同源结构域蛋白质在植物、动物和菌类中都具有广泛的保守性。