Effective method for physical mapping the DNA molecule

The method of DNA molecules physical mapping based on the algorithms of discrete optimization and graph theory was proposed. The input information consisted of the sizes of single and double restrictions fragments and the level of their measurement errors. The method presents possibilities for optimal planning of experiments and step by step construction of physical maps. Efficiency of the method and examples of its application are discussed.     

Enteroviral and Mumps Meningitis in Toronto, 1963

Virological investigations of 115 children with the aseptic meningitis syndrome during 1963 resulted in the isolation of enteroviruses from cerebrospinal fluid (CSF) and/or feces of 21 of 48 children who had no association with mumps. For the third successive year, Echo 9 was the dominant enterovirus in cases of aseptic meningitis in Toronto children, but no rashes were associated with Echo 9 meningitis during 1963, in contradistinction to previous years. Mumps virus was isolated from CSF of 25 patients by inoculation of rhesus monkey kidney cultures, and rising or elevated mumps antihemagglutinin titres in paired sera from a further 33 cases provided laboratory evidence of infection with mumps virus in 58 of 67 patients with mumps meningoencephalitis. No enlargement of salivary glands was noted in 20 laboratory-proved cases of mumps meningoencephalitis. Enteroviral meningitis occurred principally during summer, but the peak of mumps meningoencephalitis occurred during late winter.     

Linguistics of nucleotide sequences. I: The significance of deviations from mean statistical characteristics and prediction of the frequencies of occurrence of words

Mathematical models of the generation of genetic texts appeared simultaneously with the first sequencing DNA. They are used to establish functional and evolutionary relations between genetic texts, to predict the number and distribution of specific sites in a sequence and to identify "meaningful" words. The present paper deals with two problems: 1) The significance of deviations from the mean statistical characteristics in a genetic text. Anyone who has addressed himself to the statistical analysis of sequenced DNA is familiar with the question: what deviations from the expected frequencies of occurrence of particular words testify to the "biological" significance of those words? We propose a formula for the variance of the number of word's occurrences in the text, with allowance for word overlaps, making it possible to assess the significance of the deviations from the expected statistical characteristics. 2) A new method for predicting the frequencies of occurrence of particular words in a genetic text using the statistical characteristics of "spaced" L-grams. The method can be used for predicting the number of restriction sites in human DNA and in planning experiments on the physical mapping and sequencing of the human genome.     

Comparative effects of trichothecene mycotoxins on bovine platelet function: Acetyl T-2 toxin,a more potent inhibitor than T-2 toxin

The effects of the trichothecene mycotoxins (acetyl T-2 toxin, T-2 toxin, HT-2 toxin, palmityl T-2 toxin, diacetoxyscirpenol (DAS), deoxynivalenol (DON), and T-2 tetraol) on bovine platelet function were examined in homologous plasma stimulated with platelet activating factor (PAF). The mycotoxins inhibited platelet function with the following order of potency: acetyl T-2 toxin > palmityl T-2 toxin = DAS > HT-2 toxin = T-2 toxin. While T-2 tetraol was completely ineffective as an inhibitor, DON exhibited minimal inhibitory activity at concentrations above 10×10^?4M. The stability of the platelet aggregates formed was significantly reduced in all mycotoxin treated platelets compared to that of the untreated PAF controls. It is suggested that the increased sensitivity of PAF stimulated bovine platelets to the more lipophilic mycotoxins may be related to their more efficient partitioning into the platelet membrane compared to the more hydrophilic compounds.     

N-terminal Protein Processing: A Comparative Proteogenomic Analysis

N-terminal methionine excision (NME) and N-terminal acetylation (NTA) are two of the most common protein post-translational modifications. NME is a universally conserved activity and a highly specific mechanism across all life forms. NTA is very common in eukaryotes but occurs rarely in prokaryotes. By analyzing data sets from yeast, mammals and bacteria (including 112 million spectra from 57 bacterial species), the largest comparative proteogenomics study to date, it is shown that previous assumptions/perceptions about the specificity and purposes of NME are not entirely correct. Although NME, through the universal enzymatic specificity of the methionine aminopeptidases, results in the removal of the initiator Met in proteins when the second residue is Gly, Ala, Ser, Cys, Thr, Pro, or Val, the comparative genomic analyses suggest that this specificity may vary modestly in some organisms. In addition, the functional role of NME may be primarily to expose Ala and Ser rather than all seven of these residues. Although any of this group provide “stabilizing” N termini in the N-end rule, and de facto leave the remaining 13 amino acid types that are classed as “destabilizing” (in higher eukaryotes) protected by the initiator Met, the conservation of NME-substrate proteins through evolution suggests that the other five are not crucially important for proteins with these residues in the second position. They are apparently merely inconsequential players (their function is not affected by NME) that become exposed because their side chains are smaller or comparable to those of Ala and Ser. The importance of exposing mainly two amino acids at the N terminus, i.e. Ala and Ser, is unclear but may be related to NTA or other post-translational modifications. In this regard, these analyses also reveal that NTA is more prevalent in some prokaryotes than previously appreciated.Although methionine is used to initiate protein synthesis for essentially all proteins, it is subsequently removed in a large percentage of cases, either by cleavage of an N-terminal “signal ” peptide (as part of cellular translocation mechanisms or precursor activations) or by the action of specific methionine aminopeptidases (MetAPs). Approximately two-thirds of the proteins in any proteome are potential substrates for the latter N-terminal methionine excision (NME),¹ and MetAPs appear in all organisms from bacteria to eukaryotes (1). The second, or P2, amino acid in protein substrates is crucially important for NME because MetAP specificity mainly depends on the nature of this residue, a selectivity that is conserved across all species (1–5). These enzymes generally excise the N-terminal Met when the second residue is Gly, Ala, Ser, Thr, Cys, Pro, or Val (3, 6, 7), which are the amino acids smallest in size (based on radius of gyration of the side chain (8)). NME is a necessary process for proper cell functioning; it is included in the minimal genome set of eubacteria (9). Eukaryotes contain two MetAPs derived from a version in bacteria (MetAP1), and another found in archea (MetAP2) (11). Just as the deletion of MetAP eubacteria is lethal, the deletion of both MetAPs in yeast is also lethal (10).In 1988, Arfin and Bradshaw (2) observed that the specificity of NME coincided with that of the N-end rule (NER) (12, 13), a ubiquitin-dependent protein degradation process that is based on the recognition of N-terminal residues. The stabilizing residues for the NER include Gly, Ala, Ser, Cys, Thr, Pro, and Val and, with the exception of Met, the destabilizing residues are all found to be in the class of P2-residues that are not substrates for the MetAPs. This suggested that NME acts to release Met from proteins whose stability is unaffected by the NER creating at the same time a second class of proteins, who have the potential for regulated turnover downstream of the cotranslational processing, when, and if, the N-terminal Met is subsequently removed by a mechanism other than the cotranslational action of the MetAPs. However, despite extensive studies, this type of programmed protein turnover (requiring downstream removal of Met) has not been demonstrated to occur. An implication of this correlation is that exposing of the stabilizing residues may also contribute to increasing their lifetime.The stabilizing residues exposed by the action of the MetAPs can be further modified. The most extensive of these reactions is N-terminal acetylation (NTA), which can occur on as much as 70–80% of the mass of the soluble protein in eukaryotes. Although the specificity of the N-acetyltransferase (NAT) responsible is not as rigid as the MetAPs, the principal substrates in the stabilizing class are usually the four smallest residues (Gly, Ala, Ser, and Thr) (6, 14). A second class of NATs can also modify the retained Met when the adjacent residues are Asp, Glu or Asn (15). The functional importance of this modification (in either case) is not known although it has been suggested that it may exert a protective effect against spurious aminopeptidase cleavages. Recently, Hwang et al. (16) have extended the NER to include Nα-acetylated termini as also destabilizing thus providing another possible function for this modification. In contrast, to date, very few instances of Nα-acetylation have been observed in bacteria. Other modifications can also occur in both eukaryotes and prokaryotes although they are generally much more limited in scope.The specificity of the MetAPs suggest an apparent connection between NME and protein degradation. However, this connection has never been examined using high-throughput mass spectrometric data or a comparative genomics approach; thus it remains unclear whether exposing these stabilizing residues contributes to increasing protein half-life and thus represents a primary purpose of NME. (The connection between NME and NER in bacteria, which has an NER with a somewhat different profile (17), is even more obscure.) Recent studies provide some examples where disruption of NME via a single-residue substitution in the P2 position causes protein degradation (18–20); however, some of these experimental results are in conflict with the NER (13). Giglione et al. (20) have shown that NME triggers degradation of D2 protein in Caenorhabditis reinhardtii in the PSII complex after replacing the second (stabilizing) Thr residue by another amino acid to prevent NME. This replacement results in early degradation of D2 and instability of the PSII complex. From this, Giglione et al. (20) postulated that NME determines protein life-span via currently unknown machinery. However, because Bachmair et al. (12) classified Met as a stabilizing residue, it is not entirely clear why substituting one stabilizing residue (Met) by another one (Gly, Ala, Ser, Cys, Thr, Pro, or Val) should affect protein stability and the substitution may have other deleterious effects that are manifested in different ways.The logic for analyzing NME and NER is shown in Fig. 1. NME exposes 7 different residues as new N termini of proteins. The natural conclusion that has become a dogma of NME is that these seven residues are exposed for a functional reason. The broad scope of NME suggests a universal reason that surpasses any particular protein''s role. In turn the comparative genomics postulate (function suggests conservation) leads to the conclusion that the seven residues should be evolutionarily conserved at position P2 of proteins. However, because only two out of the seven residues are conserved, we argue that one of the two assumptions in Fig. 1A must be incorrect and put forth the alternative logic depicted in Fig. 1B, which matches our analysis across dozens of species. According to this logic, NME accomplishes the goal of exposing Ala and Ser by exposing all residues with side chains smaller or comparable in size to Ala and Ser (G, T, V, P, and C). These residues are thus inconsequential players that are not functionally important (and are not evolutionarily conserved) at P2.Open in a separate windowFig. 1.Two alternative cases for NME function. A, NME exposes seven residues to be new N termini of proteins. Because this is presumably for some functional reason, the conventional assumption is that all seven residues must have functional importance as N termini. By the comparative genomics postulate (as defined in the text), evolutionary conservation of all seven at P2 should be observed. If all of these residues are not conserved, one of the two assumptions must be incorrect; either not all seven residues are important or the comparative genomics postulate is invalid. B, Given that the comparative genomics postulate holds, and only two of the seven residues are of functional importance as N termini, then the other five residues are inconsequential players and only these two residues should be evolutionarily conserved.In this report, we examine the connection between the specificity of NME and stabilizing residues of NER. In doing so, data sets from bacteria (including 112 million mass spectrometric spectra from 57 species), yeast, and mammals, were analyzed for N-terminal peptides both with respect to the excision (or not) of initiator Met residues and the distribution of P2-residues. The results reveal a strong preference of Ala and Ser as P2-residues. However, this process does not appear to be linked to the NER other than being generally compatible with it. These studies also demonstrate a much greater than expected number of Nα-acetylation events in some bacteria.     

Inflammatory pre-conditioning of mesenchymal multipotent stromal cells improves their immunomodulatory potency in acute pyelonephritis in rats

Background aimsAcute pyelonephritis is one of the most frequent infectious diseases of the urinary tract and a leading cause of kidney failure worldwide. One strategy for modulating excessive inflammatory responses in pyelonephritis is administration of mesenchymal multipotent stromal cells (MMSCs).MethodsThe putative protective effect of injection of MMSCs against experimental acute pyelonephritis was examined. We used in vivo experimental model of APN where bacteria are introduced in the bladder of rat. Three days after, intravenous injection of MMSCs was done. On the 7th day blood samples and kidneys were taken for further analysis.ResultsWe found obvious signs of oxidative stress and inflammation in the kidney in acute pyelonephritis in rats. Particularly, pro-inflammatory cytokine tumor necrosis factor-α levels, malondialdehyde, nitrite and myeloperoxidase activity were significantly increased. Histologic evaluation revealed numerous attributes of inflammation and tissue damage in the kidney. Treatment with MMSCs caused a remarkable decrease of all of these pathologic signs in renal tissue. Also, activated leukocytes induced pre-conditioning-like signaling in MMSCs. We showed alterations of expression or activity of inducible nitric oxide synthase, transforming growth factor-β, matrix metalloproteinase-2 and glycogen synthase kinase-3β, which could mediate immunomodulation and protective effects of MMSCs. This signaling could be characterized as inflammatory pre-conditioning.ConclusionsThe beneficial capacity of MMSCs to alleviate renal inflammation was more pronounced when pre-conditioned MMSCs were used. This approach could be used to prime MMSCs with different inflammatory modulators to enhance their engraftment and function in an immunoprotected fashion.     

吐鲁番市胃食管反流病发生相关危险因素分析（附280例报道）

目的：探讨280例胃食管反流病（GERD）的分布特点及危险因素。方法：对临床诊断和胃镜确诊的280例GERD患者进行临床和风险因子相关性分析。结果：不论汉族还是维族,男性患者比例均明显高于女性;汉族患者高发年龄段早于维族患者（z=-2．939,P=0．003,）;汉族和维族患者占反流性食管炎和Barrett食管比例分别为42．4％、81_3％及56．5％、18．8％,其中汉族患者Barrett食管比例较高（X2=14．358,P=0．000）;肥胖、习惯性便秘、重体力活动者、饮食习惯不良在维族患者中的比例较高（P〈0．001）。结论：GERD与性别、年龄密切相关,男性多于女性,汉族患者发病年龄高峰旱于维族患者;汉族患者Barrett食管发生比例高于维族患者;肥胖、习惯性便秘、重体力活动、饮食习惯不良可能是GERD尤其是维族人群GERD的危险因素。