Retinal Attachment Instability Is Diversified among Mammalian Melanopsins

Melanopsins play a key role in non-visual photoreception in mammals. Their close phylogenetic relationship to the photopigments in invertebrate visual cells suggests they have evolved to acquire molecular characteristics that are more suited for their non-visual functions. Here we set out to identify such characteristics by comparing the molecular properties of mammalian melanopsin to those of invertebrate melanopsin and visual pigment. Our data show that the Schiff base linking the chromophore retinal to the protein is more susceptive to spontaneous cleavage in mammalian melanopsins. We also find this stability is highly diversified between mammalian species, being particularly unstable for human melanopsin. Through mutagenesis analyses, we find that this diversified stability is mainly due to parallel amino acid substitutions in extracellular regions. We propose that the different stability of the retinal attachment in melanopsins may contribute to functional tuning of non-visual photoreception in mammals.     

Cloning of a Vero toxin (VT2) gene from a VT2-converting phage isolated from Escherichia coli 0157: H7

Abstract A Vero toxin (VT2 or Shiga-like toxin II)-converting phage was isolated from Escherichia coli 0157: H7 strain J-2. Nontoxigenic E. coli C600 produced VT2 when lysogenized with the toxin-converting phage. Eco RI fragments of the phage DNA were ligated with Eco RI-digested pBR322 or pUC118 and were transformed into E. coli MC1061 or MV1184. Transformants exhibiting VT2 production commonly contained a 4.6 kb Eco RI fragment. It was found that a 2.3 kb Kpn I- Sph I fragment coded VT2 production and that this fragment hybridized weakly with the 2.1 kb fragment encoding VT1.     

Suppressors of temperature-sensitive mutations in a ribosomal protein gene, rpsL (S12), of Escherichia coli K12

Summary Temperature-sensitive (ts) mutations were isolated within a ribosomal protein gene (rpsL) of Escherichia coli K12. Mutations were mapped by complementation using various transducing phages and plasmids carrying the rpsL gene, having either a normal or a defective promoter for the rpsL operon. One of these mutations, ts118, resulted in a mutant S12 protein which behaved differently from the wild-type S12 on CM-cellulose column chromatography. Suppressors of these ts mutations were isolated and characterized; one was found to be a mutation of a nonribosomal protein gene which was closely linked to the RNAase III gene on the E. coli chromosome. This suppressor, which was recessive to its wild-type allele, was cloned into a transducing phage and mapped finely. A series of cold-sensitive mutations, affecting the assembly of ribosomes at 20°C, was isolated within the purL to nadB region of the E. coli chromosome and one group, named rbaA, mapped at the same locus as the suppressor mutation, showing close linkage to the RNAase III gene.     

Carboxyl Terminal Segment of T4 Gene 32 Protein is Dispensable in Vivo

We obtained a mutant of bacteriophage T4 which overcame thedeficiency in gene 49 endonuclease. The new mutation occurredin gene 32 and the mutant, which was viable, produced an amberfragment under non-suppressed conditions, lacking about 30 aminoacid residues at the carboxyl terminus. Its growth, recombination,and resistance to UV irradiation were affected to various degreesby the particular suppressor tRNA present. Growth was increasedby Su2⁺ to nearly that of the wild type, but growth of all otherswas reduced in the presence and absence of suppressors, suggestingthat the terminal domain of gene 32 protein is not indispensablefor the function but modulates it. We discuss the mechanismby which the mutation overcomes the defect in gene 49 endonuclease. ¹ This paper is dedicated to the memory of the late Dr. JojiAshida. (Received November 22, 1982; Accepted February 21, 1983)     

Electron microscopical studies on the thyroid of a cyclostome,Lampetra japonica,during its upstream migration

Summary Electron microscopical studies were made of the thyroid gland of an adult lamprey, Lampetra japonica, in the upstream migration period.The thyroid consists of many usual follicles containing the colloid in their lumina, and a large parafollicle without colloid. The paper concerns only the usual follicle.The follicle cells found in the usual follicle wall are classified into three types; 1. a non-ciliated taller cell, 2. a ciliated taller one, and 3. a non-ciliated cuboidal one. From their cytoplasmic fine structure, it is considered that all these cells are essentially identical and differences among them are due to their functional state.All these type cells are characterized by irregularly developed interdigitations and aggregates of tonofilaments throughout the cytoplasm, especially in the perinuclear region. Although the rough-surfaced endoplasmic reticulum and the Golgi apparatus are fairly well developed in the first and second type cells, the cisternae are not so large-vacuolated but flattened, and the cytoplasm is more compact as compared with that of the higher vertebrate. In the third type cell, the cytomembranes are poorly developed.Large dense inclusion-bodies consisting of heterogeneously dense materials, of lamellar structures, and of less dense vacuoles, which are found often in taller follicle cells, are also characteristic for the lamprey thyroid. The body which might be intimately related to the Golgi apparatus is considered to be a kind of lysosomes and it perhaps corresponds to the yellow pigment observed by light microscopy.In the apical part of the cytoplasm in taller cells, there are three kinds of granules or vesicles; numerous small vesicles considered to be derived from the Golgi apparatus, a few small dense granules which seem to originate from the Golgi region, and a few large less-dense granules.In the third type cell, the cytomembranes are not so well developed as those of the first and second type cells. The large heterogeneously dense bodies and the cytoplasmic granules are very few in number.Around the follicle of the lamprey thyroid, there are a dense basement membrane and a relatively compact connective tissue with few blood capillaries. Characteristic fat cells are found in the connective tissue.     

Analysis of Vibrio cholerae O139 Bengal Isolated from Different Geographical Areas Using Macrorestriction DNA Analysis

Vibrio cholerae O139 isolated from different countries, as well as from different locations within a country, were examined using macrorestriction DNA analysis to determine the clonality of the O139 strains. NotI digests of genomic DNA of representative strains from Nepal, India, Bangladesh, China, Thailand, and Malaysia revealed very similar but not identical patterns. Examinations of the banding patterns generated by pulsed-field gel electrophoresis of strains isolated within countries revealed complete homogeneity. These results further reiterate the spread of an identical clone of V. cholerae O139 although it appears that genetic polymorphism among the O139 strains is becoming apparent.     

Typing of Verotoxins by DNA Colony Hybridization with Poly- and Oligonucleotide Probes,a Bead-Enzyme-Linked Immunosorbent Assay,and Polymerase Chain Reaction

To identify the type of Verotoxins (VT) produced by Verocytotoxin-producing Escherichia coli (VTEC), a sensitive bead-enzyme-linked immunosorbent assay and polymerase chain reaction with common and specific primers to various VTs (VT1, VT2, VT2vha, VT2vhb, and VT2vp1) were developed. Together with colony hybridization tests with oligo- and polynucleotide probes, these methods were applied to VTEC isolates to type the VT produced. The toxin types of 26 of 37 strains were identified, but the reaction profiles in assays of the remaining 11 strains suggested the existence of new VT2 variants. The application of these identification procedures may be useful as a tool for clinical and epidemiological studies of VTEC infection.