Raman spectroscopic analysis of the effect of ultraviolet irradiation on calf thymus DNA

Raman spectroscopy was used for the first time to detect the effect of independent UVA (ultraviolet-A: 320-400nm) and UVB (ultraviolet-B: 280-320 nm) irradiation on the calf thymus DNA in aqueous solution. After both UVA and UVB irradiation for 1h or 3h, the damage to the conformation of DNA was moderate, but the reduction of the B-form DNA component was obvious. Both UVA and UVB caused significant damage to the deoxyribose moiety and bases, among which the pyrimidine base pairs were more seriously affected. There appeared to be preferential damaging sites on DNA molecules caused by UVA and UVB irradiation. UVA irradiation caused more damage to the deoxyribose than UVB irradiation, while UVB irradiation caused more significant damage to the pyrimidine moiety than UVA irradiation. After UVB irradiation for 3h, unstacking of the AT base pairs and the cytosine ring took place, severe damage to the thymine moiety occurred, and some base pairs were modified. Moreover, with either UVA or UVB irradiation for 3h,the photoreactivation of DNA occurred. The damage to the DNA caused by UVB was immediate, while the damage caused by UVA was proportional to the irradiation duration. The experimental results partly indicate the formation of some cyclobutane pyrimidine dimers and (6-4) photoproducts.     

Immunity repressor of bacteriophage P2: Identification and DNA-binding activity

The product of gene C of the temperate bacteriophage P2, the immunity repressor, can be detected as a unique band eluting from phosphocellulose columns at 0.12 m-potassium phosphate when differentially labelled with a radioactive amino acid: the band is absent when phages that either have lost gene C through deletion or carry a suppressor-sensitive mutation in the gene are used. The repressor in its monomeric form is about 11,000 in molecular weight. At near physiological salt concentrations, the form predominantly recovered is the dimer.In filter-binding assays, the partially purified repressor binds wild-type P2 DNA strongly. It does not bind DNA of P2 vir94, a deletion that removes all the genetic elements involved in the regulation of lysogeny; it also does not bind, or binds inefficiently, DNA of P2 vir3, a mutation in the operator that controls the early replicative functions of P2. At the concentrations employed, the dimer is the active form in binding.The P2 repressor clearly differs in several features from the well-studied immunity repressor of bacteriophage lambda.     

Molecular characterization and phylogenetic position of a new mariner-like element in the coastal crab, Pachygrapsus marmoratus

Mariner-like elements (MLEs) are class-II transposable elements that move within the genome of their hosts by means of a DNA-mediated "cut and paste" mechanism. MLEs have been identified in several organisms, from most of the phyla. Nevertheless, only a few of the sequences characterized contain an intact open reading frame. Investigation of the genome of a coastal crab, Pachygrapsus marmoratus, has identified nine Pacmmar elements, two of which have an open reading frame encoding a putatively functional transposase. Nucleic acid analyses and comparison with the previous data showed that the GC contents of MLEs derived from coastal organisms such as P. marmoratus are significantly higher than those of terrestrial MLEs and significantly lower than those of hydrothermal ones. Furthermore, molecular phylogeny analyses have shown that Pacmmar elements constitute a new lineage of the irritans subfamily within the mariner family.     

Cloning of both ends and the thermo-inducible genes A and B of bacteriophage Mu on a multicopy plasmid

Employing the recombinant DNA technique, two hybrid plasmids were constructed carrying both ends of bacteriophage Mu DNA in two different orientations. The expression of the early Mu genes located on these plasmids is thermo-inducible.     

Distribution and evolution of short tandem repeats in closely related bacterial genomes

Simultaneous identification and comparison of perfect and imperfect microsatellites within a genome is a valuable tool both to overcome the lack of a consensus definition of SSRs and to assess repeat history. Detailed analysis of the overall distribution of perfect and imperfect microsatellites in closely related bacterial taxa is expected to give new insight into the evolution of prokaryotic genomes. We have performed a genome-wide analysis of microsatellite distribution in four Escherichia coli and seven Chlamydial strains. Chlamydial strains generally have a higher density of SSRs and show greater intra-group differences of SSR distribution patterns than E. coli genomes. In most investigated genomes the distribution of the total lengths of matching perfect and imperfect trinucleotide repeats are highly similar, with the notable exception of C. muridarum. Closely related strains show more similar repeat distribution patterns than strains separated by a longer divergence time. The discrepancy between the preferred classes of perfect and imperfect repeats in C. muridarum implies accelerated evolution of SSRs in this particular strain. Our results suggest that microsatellites, although considerably less abundant than in eukaryotic genomes, may nevertheless play an important role in the evolution of prokaryotic genomes and several gene families.     

Comparative study of the L1 family in the genus Mus. Possible role of retroposition and conversion events in its concerted evolution

The long interspersed repetitive family L1 was analysed in different species belonging to the genus Mus. It is shown to be highly conserved even in M.n. setulosus, which diverged from the other species around ten million years ago. The study of the linkage between diagnostic restriction sites in the various species and the sequence variations of different regions of the L1Md repeat shows that the L1 family undergoes concerted changes involving subsets of repeats. The rate at which this homogenization process occurs does not appear to be the same for all the subfamilies detected. The L1Md repeat in the twelfth intron of the serum albumin gene of Balb/c mice is shown to be a recent insertion. The role retroposon- and gene conversion-like events may play in the concerted evolution of the L1 family is discussed.     

Identification and mapping of a 60 bp EcoRI fragment in the Dictyostelium discoideum ribosomal DNA

We have re-examined the organization of the transcribed sequences in the ribosomal DNA repeat unit of Dictyostelium discoideum. In addition to the four EcoRI fragments previously reported, we have identified and cloned a fifth fragment which defines a small portion of the 25S ribosomal RNA. The fragment is 60 bp long and is located between the 1.5 kbp and the 3 kbp EcoRI fragments of the ribosomal DNA repeat unit.     

紫外线B区对小牛胸腺DNA损伤的拉曼光谱研究

用拉曼光谱检测了远紫外区UVB(280m～320nm)对小牛胸腺DNA的损伤,并对这个区段紫外线对DNA的不同照射时间时的损伤特征进行了比较分析。实验中所用的紫外线强度与太阳光强度相当。结果表明,辐照3h以内时,UVB对DNA的构型有较明显的损伤,这可能是受到嘧啶碱基损伤的影响。相对而言,UVB对脱氧核糖和碱基的损伤要严厉得多。就碱基对的受损伤程度来说,嘧啶碱受损最严重,部分证明了环丁烷嘧啶二聚体和6-4光产物的形成。经过3h UVB照射,AT碱基对和和胞嘧啶环的堆叠程度有所瓦解,一些碱基对受到修饰。而一些拉曼特征峰强度的反复波动,则说明了长时间的紫外照射可以导致部分DNA光复活的产生。进一步分析发现,UVB以一种较快的方式对DNA产生损伤。     

The alpha and beta subunits of lamb kidney Na,K-ATPase are both glycoproteins

We have determined the carbohydrate compositions of the protein components of lamb kidney Na,K-ATPase. The α subunit contains a total of about 16 monosaccharide residues per mol of protein, while the β subunit contains about 36 residues per mol. The γ protein, a proteolipid associated with the Na,K-ATPase, contains only traces of carbohydrate. A comparison of our results with those of others shows considerable variability in the carbohydrate compositions of α and β subunits from different species.     

Genetic reconstruction and functional analysis of the repeating lipoyl domains in the pyruvate dehydrogenase multienzyme complex of Escherichia coli

The dihydrolipoamide acetyltransferase component (E2p) of the pyruvate dehydrogenase complex of Escherichia coli contains three highly homologous sequences of about 100 residues that are tandemly repeated to form the N-terminal half of the polypeptide chain. All three sequences include a lysine residue that is a site for lipoylation and they appear to form independently folded functional domains. These lipoyl domains are in turn linked to a much larger (about 300 residues) subunit-binding domain of the E2p chain that aggregates to form the octahedral inner core of the complex and also contains the acetyltransferase active site. In order to investigate whether individual lipoyl domains play different parts in the enzymic mechanism, selective deletions were made in vitro in the dihydrolipoamide acetyltransferase gene (aceF) so as to excise one or two of the repeating sequences. This was facilitated by the high degree of homology in these sequences, which allowed the creation of hybrid lipoyl domains that closely resemble the originals. Pyruvate dehydrogenase complexes incorporating these genetically reconstructed E2p components were purified and their structures were confirmed. It was found that the overall catalytic activity, the system of active site coupling, and the ability to complement pyruvate dehydrogenase complex mutants, were not significantly affected by the loss of one or even two lipoyl domains per E2p chain. No special role can be attached thus far to individual lipoyl domains. On the other hand, certain genetic deletions affecting the acetyltransferase domain caused inactivation of the complex, highlighting particularly sensitive areas of that part of the E2p chain.     

Population genetic structure of Penaeus monodon, in relation to monsoon current patterns in Southwest, East and Andaman coastal waters of India

The black tiger shrimp (Penaeus monodon), a commercially important penaeid species, is widely distributed across the Indo-Pacific region. Genetic diversity in P. monodon collected from eight geographical regions in Southwest, East and Andaman coastal waters of India (N = 418) was investigated using 10 polymorphic microsatellite loci. Average observed heterozygosity at sampled loci were high, ranging from 0.643 (Coromandel Coast) to 0.753 (South Andaman). Pairwise F_ST (ranged from 0.005 to 0.078) and R_ST (ranged from 0.005 to 0.171) estimates revealed surprisingly strong and statistically significant genetic structure among tiger shrimp populations. A synthetic map generated by multidimensional scaling shows an apparent cline in allele frequencies paralleling the roughly circular flow of surface currents in the Bay of Bengal. Significant heterozygote deficiencies were noted in most population samples at most loci. Andaman Island sites showed the highest diversity. Recognition of high genetic diversity and distinct population structuring of P. monodon in Indian seas has important implications for future domestication of this species in India, for two reasons: identification of the best wild founding stocks for aquaculture and, subsequently, the potential impacts of release of domesticates to the wild, either accidentally or deliberately (i.e. for stock enhancement).     

Conserved function of Rho-related Rop/RAC GTPase signaling in regulation of cell polarity in Physcomitrella patens

Cell polarity is fundamentally important to growth and development in higher plants, from pollen tubes to root hairs. Basal land plants (mosses and ferns) also have cell polarity, developing protonemal apical cells that show polar tip growth. Flowering plants have a distinct group of Rho GTPases that regulate polarity in polarized cell growth. Rop/RAC signaling module components have been identified in non-flowering plants, but their roles remain unclear. To understand the importance and evolution of Rop/RAC signaling in polarity regulation in land plants, we examined the functions of PpRop and PpRopGEF in protonemal apical cells of the moss Physcomitrella patens. Inducible overexpression of PpRop2 or PpRopGEF3 caused depolarized growth of tip-growing apical cells. PpRop2 overexpression also caused aberrant cross wall formation. Fluorescent protein-tagged PpRop2 localized to the plasma membrane, including the cross wall membrane, and fluorescent-tagged PpRopGEF3 showed polarized localization to the tip region in apical cells. Thus, our results suggest common functions of PpRop and PpRopGEF in the tip-growing apical cells and the importance of a conserved Rop/RAC signaling module in the control of cell polarity in land plants.     

An effect of ATP on the permeability of transport-competent plasma membrane vesicles from mouse fibroblasts.

Methods for predicting peptide chain conformation have been applied to amino acid sequences adjacant to the carbohydrate attachment sites of glycoproteins containing the $\text{N}$-glycosylamine type of protein-carbohydrate linkage. Of 31 glycosylated residues examined 30 occur in sequences favouring turn or loop structures. Twentytwo of the glycosylated asparagine residues occur in tetrapeptides predicted to have the β-turn conformation. Carbohydrate attachment is therefore associated with peptide sequences which favour the formation of β-turn or other turn or loop structures.     

Identification of a human clustered G + C-rich DNA family of repeats (Sau3A family)

Sau3A digestion of human G + C-rich DNA molecules yields discrete bands of approximately 70 and 140 base-pairs, under-represented in A + T-rich DNA molecules and in total DNA. We have cloned the 70 base-pair band in a plasmid vector and isolated a representative recombinant clone that identifies a new human family of repeats, the Sau3A family. The new family has been characterized for a number of parameters: genomic organization; reiteration frequency; sequence analysis; and distribution in a human genomic library. The Sau3A sequence (68 base-pairs in length, 53% G + C) is present in approximately 4 X 10(4) copies/haploid genome; the family is characterized by a cluster organization and is confined to a limited fraction (0.5%) of phages of a human genomic library. Southern blot hybridizations of the cloned sequence to restriction digests of total human DNA and of isolated genomic clones does not show the involvement of Sau3A blocks in long-range periodicities for any of the enzymes tested. The data suggest either a high sequence variability in the family or a complex organization of Sau3A sequence domains.