The structure and biochemistry of the vacuolar H+ ATPase in proximal and distal urinary acidification

Vacuolar H⁺ ATPases participate in renal hydrogen ion secretion in both the proximal and distal nephron. These plasma membrane forms of the vacuolar H⁺ ATPase are regulated physiologically to maintain the acid-base balance of the organism. Proton transporting renal cells have requirements for constitutive acidification of intracellular compartments for normal endocytic and secretory functions. Recent experiments have begun to reveal how the kidney regulates these proton pumps independently. Vacuolar H⁺ ATPases are a family of structurally similar enzyme which differ in the composition of specific subunits. Cytosolic regulatory enzymes are present in renal cells which may affect vacuolar H⁺ ATPases in certain membrane compartments selectively. The vacuolar H⁺ ATPase in the plasma membrane of intercalated cells resides in a specialized proton-transporting apparatus that translocates the enzyme between an intracellular membrane pool and the plasma membrane in response to physiologic stimuli.This review will focus on the structure, enzymology, and regulation of the vacuolar H⁺ ATPase in the mammalian kidney. Because of space limitations, it will cover predominantly work from our laboratory. However, a number of investigators, including Brown (Brownet al., 1987, 1988a,b, 1989), Burckhardt (Sabolicet al., 1985; Turriniet al., 1989; Simon and Burckhardt, 1990), Madsen and Tisher (Madsen and Tisher, 1985; Verlanderet al., 1987, 1989). Steinmetz (Steinmetz, 1986; Stetson and Steinmetz, 1986), Schwartz (Scwartzet al., 1985, 1988; Satlin and Schwartz, 1989), Sabatini and Kurtzman (Sabatiniet al., 1990a,b), DuBose (Diaz-Diazet al., 1986; Gurich and DuBose, 1989), Al-Awqati (Van Adelsberg and Al-Awqati, 1986), and their coworkers, and many other investigators have made important contributions to this field.     

CFTR: Domains, Structure, and Function

Mutations in the gene encoding the cystic fibrosis transmembrane conductance regulator (CFTR) cause cystic fibrosis (CF) (Collins, 1992). Over 500 naturally occurring mutations have been identified in CF gene which are located in all of the domains of the protein (Kerem et al., 1990; Mercier et al., 1993; Ghanem et al., 1994; Fanen et al., 1992; Ferec et al., 1992; Cutting et al., 1990). Early studies by several investigators characterized CFTR as a chloride channel (Anderson et al.; 1991b,c; Bear et al., 1991). The complex secondary structure of the protein suggested that CFTR might possess other functions in addition to being a chloride channel. Studies have established that the CFTR functions not only as a chloride channel but is indeed a regulator of sodium channels (Stutts et al., 1995), outwardly rectifying chloride channels (ORCC) (Gray et al., 1989; Garber et al., 1992; Egan et al., 1992; Hwang et al., 1989; Schwiebert et al., 1995) and also the transport of ATP (Schwiebert et al., 1995; Reisin et al., 1994). This mini-review deals with the studies which elucidate the functions of the various domains of CFTR, namely the transmembrane domains, TMD1 and TMD2, the two cytoplasmic nucleotide binding domains, NBD1 and NBD2, and the regulatory, R, domain.     

Non-random association of human acrocentric chromosomes

Summary The association of human acrocentric chromosomes was found to be nonrandom using fluorescence technic.The occurrence of chromosomes D13-15 in Robertsonian rearrangement has been shown to be non-random (Hecht et al., 1968). Ohno et al. (1961) have postulated these rearrangements might be related to acrocentric association. In two studies (Nakagame, 1969; Shaw et al., 1969), however, a random distribution of the D group chromosomes in association was found by autoradiography in lymphocytes. The recently described fluorescence technic (Caspersson et al., 1971) permits a reliable identification of each of the 23 pairs and we would like to report the results of our re-examination of acrocentric association by this method.

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Zusammenfassung Die Assoziation menschlicher akrozentrischer Chromosomen hat sich als nicht zufällig erwiesen. Es wurde die Fluorescenztechnik benutzt.

     

Structure and affinities of the petrified plants from the Cretaceous of Japan and Saghalien,V. Tree fern stems from Hokkaido,Paracyathocaulis ogurae gen. et comb. nov. andCyathocaulis yezopteroides sp. nov.

Paracyathocaulis ogurae gen. et comb. nov. andCyathocaulis yezopteroides sp. nov. are Late Cretaceous tree-fern stems with cyatheaceous affinity. They are comparable with the Recent scaly cyatheoids (Cyathea s.l.), as well as with the Cretaceous and Tertiary fossil stems that have similar anatomy. All of these tree ferns, including the Late Jurassic to Early CretaceousOguracaulis Tidwellet al., comprise a well-defined morphological complex within Cyatheaceae, and are considered to be monophyletic. The taxonomy of this complex is analyzed cladistically, and the cladogram is phylogenetically evaluated by reference to the chronological data of fossils.     

Books on Insect Biodiversity and Conservation

Reviews are provided for four books relevant to the studies on the biodiversity conservation of insects: Insect diversity conservation (Samways 2005), Insect conservation biology (Stewart et al. 2007), Ecology of insects: concepts and applications (Speight et al. 2008, 2nd edn), and Insect conservation: a handbook of approaches and methods (Samways et al. 2009).     

On the validization of the name Chironomus (Camptochironomus) pallidivittatus sensu edwards, 1929 and elimination of the name Chironomus pallidivittatus Malloch, 1915

The name Chironomus pallidivittatus sensu Edwards is widely used by specialists for the European species described by Edwards, whereas the use of the name Ch. tentans var. pallidivittatus Malloch is limited. In the light of the wide use of the name Chironomus pallidivittatus sensu Edwards, particularly in fields outside taxonomy, combined with virtually no publications on the Ch. tentans variety pallidivittatus Malloch, we recommend that the original Malloch’s application of the name should be suppressed in favor of that of Edwards, 1929. At the same time, synonymy of Ch. pallidivittatus s. Malloch with Ch. tentans Fabricius supposed by Townes (1945) is erroneous. Detailed cytogenetic (Kiknadze et al., 1996) and morphological (Shobanov et al., 1999) studies of North American, European, and Asian populations demonstrated the common occurrence of Ch. dilutus Shobanov, Kiknadze et Butler, 1999 in the Nearctic. This species is different from Ch. tentans but identical with Ch. tentans var. pallidivittatus Malloch, 1915 and the “Nearctic Ch. tentans” sensu Townes (1945).     

Time to update the sectional classification of Ilex (Aquifoliaceae): New insights from Ilex phylogeny,morphology, and distribution

Resolving the infrageneric classification of species-rich genera has been challenging in plant taxonomy. Ilex L. is a subcosmopolitan genus with over 600 species of dioecious trees and shrubs. Many classification systems based on morphological data have been proposed during the past 250 years. However, these systems (such as Loesener's and Galle's systems) may not truly reflect Ilex's evolutionary trajectories because most of those system's infrageneric hierarchies are not monophyletic. In this study, we reconstructed a phylogeny of Ilex L. comprising 15 moderately to highly supported clades using rigorously identified samples (202 species) and closely authenticated gene sequences of three nuclear genes [internal transcribed spacer (ITS), external transcribed spacer (ETS), and nepGS]. The newly generated phylogenetic tree resembles essentially that of the nuclear tree of Manen et al., but shows conspicuous topological differences with the phylogeny of Yao et al. Closely scrutinizing morphological variation and distributional patterns of 202 species, this study found that most lineages of Ilex identified herein are well defined by a particular trait or a combination of morphological and distributional traits, displaying phylogeny–morphology–distribution conformity that has seldom been uncovered in previous studies. Given the general phylogeny–morphology–distribution conformity revealed in this genus, we put forward an updated sectional classification system for Ilex that temporarily contains 14 sections. The new classification will provide a robust framework for studying the evolution and diversification of this ecologically and economically important genus.     

Genetic analysis of the two groups of duplicated genes coding for mitochondrial malate dehydrogenase in Zea mays: Possible origin of Mdh genes by chromosome segment duplication

Summary The results presented in this paper demonstrate that there are four mitochondrial malate dehydrogenase genes (mMdh1, mMdh2, mMdh3, and mMdh4) as was proposed by Yang et al. (1977), but with the following modification. Yang et al. (1977) postulated that mMdh1 and mMdh2 were closely linked as were mMdh3 and mMdh4. They also proposed that mMdh3 and mMdh4 arose by gene duplication from mMdh1 and mMdh2 respectively. In the modified model mMdh3 and mMdh4 are linked, but the other two genes mMdh1 and mMdh2 are unlinked. In addition, we demonstrate using reversible denaturation techniques, which mMDH isozymes are heterodimeric in structure. Hence, the origin of all the mMDH isozymes has been elucidated.Since the number of genes in the modified mMDH model remains the same as the Yang et al. model (1977), the original nomenclature is used. We illustrate further that the three gene model proposed by Goodman et al. (1980) and accepted by their collaborators, Newton and Schwartz (1980) is not consistent with data from our genetic crosses. However, the modified version of the Yang et al. (1977) model will account for all MDH data published to date. The psossible role of chromosome segment duplication in the evolution of the MDH genes in Zea mays is discussed.This research was supported, in part, by NIH Grants T501-GM296, GM22733, and by the N.C. Agricultural FoundationPaper No. 6833 of the Journal Series of the North Carolina Agricultural Research Service     

The measurement of membrane potential using optical indicators

Conclusions Optical methods have become established as a major experimental protocol for following membrane potential. They can provide a rapid, continuous record of the potential and have a very wide applicability. However, when used to make quantitative assertions about membrane potential, optical methods have a number of weaknesses. Even the most reliable calibration procedures depend on accurate evaluation of a small number, namely the internal ion concentration, in a large background, that is total ion levels. However, a consensus seems to be emerging that the plasma membrane potential of non-excitable cells nevertheless has considerable magnitude: typical values are –60 mV for lymphocytes (Rinket al., 1980), –20 to –100 mV, depending on metabolic load, for Ehrlich ascites tumour cells (Philo & Eddy, 1978; but see also Smith & Robinson, 1980), and –66 to –86 mV for neutrophils (Tathamet al., 1980). In our own experiments using monolayer cultures of cells grown to confluence (Bashfordet al., 1981) the potential across the plasma membrane is of the order of –100 mV (see Fig. 2). Membrane potentials of similar magnitude have been found using ion-distribution methods and microelectrodes in neuroblastoma cells and lymphocytes (Deutschet al., 1979a,b). In the latter studies ions of different charge were used to provide upper and lower estimates of the potential, the presumed effects of binding being very different for anions and cations. A similar approach, in this case the use of optical indicators of different charge, has been taken by Rinket al. (1980), and this would seem to be one way in which to diminish the uncertainties involved in dye calibration. Unfortunately many anions, particularly oxonols, form complexes with valinomycin (Lavie & Sonenberg, 1980; Rinket al., 1980), although we have found no evidence for such a complex with bis isoxazolone oxonols (J.C. Smith and C.L. Bashford, unpublished observations). It is apparent that calibration procedures not dependent on valinomycin should be sought in order to establish optical methods as a quantitative approach to the study of membrane potential.     

Baby in the bathwater: Should we abandon the use of body temperature thresholds to quantify expression of torpor?

Boyles et al. (this issue) argue against the use of body temperature (T_b) thresholds to quantify the expression of torpor in endotherms and our purpose is to provide a counterpoint argument. We contend that T_b thresholds provide valuable information about ecological factors influencing the evolution of thermoregulation. We also point out shortcomings of the so-called heterothermy index proposed as an alternative. However, to be clear, we do agree with Boyles et al. (this issue) that the use of torpor thresholds can limit some aspects of the study of thermoregulation and applaud the more widespread incorporation of theoretical underpinnings proposed by Boyles et al. (this issue) and others.     

Role of 2-ketobutyrate as an alarmone in E. coli K12: inhibition of adenylate cyclase activity mediated by the phosphoenolpyruvate: glycose phosphotransferase transport system

Summary 2-ketobutyrate and its analogues were found to inhibit strongly and transiently the rate of -galactosidase synthesis in Escherichia coli K12. This effect was ascribed to a strong and transient inhibition of the adenylate cyclase activity. By using pts mutants, we showed, in agreement with our previous results (Daniel et al. 1983), that the likely target of 2-ketobutyrate and its analogues is the phosphoenolpyruvate: glycose phosphotransferase transport system (PTS). Furthermore, evidence for such a cascade effect caused by 2-ketobutyrate and its analogues allowed us to corroborate our previous proposal (Daniel et al. 1983) that 2-ketobutyrate, a precursor of isoleucine, acts as an E. coli alarmone monitoring the passage from anaerobic to aerobic growth conditions.     

Confounding social and mating systems predictably lead to biased results when examining the evolution of cooperative breeding in cichlids: A response to Tanaka et al.

In 2017, we demonstrated that transitions to cooperative breeding in Lamprologine cichlid fishes were not related to a species’ social mating system (Dey et al. 2017. Nature Ecology & Evolution, 1 , 137). This contrasted previous evidence that monogamy (and a low degree of promiscuity) promoted transitions to cooperative breeding in other taxa. Recently, Tanaka et al. (2018. Ethology, 124 , 777–789) critiqued our study and argued that a re‐analysis of the data shows transitions to cooperative breeding are promoted by non‐monogamous mating systems. Here, we show that Tanaka et al.'s critique contains numerous inaccuracies. In addition, we show that the results put forth by Tanaka et al. emerge only under the extreme scenario in which all cooperative breeding species are classified as non‐monogamous, which we argue arises because Tanaka et al. confound social systems and mating systems. While we agree that there is uncertainty regarding the mating system of some Lamprologine species, we argue this uncertainty was sufficiently addressed through the extensive sensitivity analyses conducted in our original study.     

The subspecies concept in primatology: The case of mountain gorillas

The criteria for the application of subspecific units in living primate populations have received little attention relative to other vertebrate taxa, even though they have important implications for conservation strategies for many nonhuman primate populations. One of the most critically endangered primates is the mountain gorilla,Gorilla gorilla beringei, of which 600 animals exist in east-central Africa. FollowingSarmiento et al. (1996), taxonomists have proposed splitting these populations into two subspecies as part of a revised taxonomy of the genusGorilla. In this paper I review the application of the subspecies concept in primatology, using the gorillas of Bwindi Impenetrable National Park and the Virungas as case studies. An examination of genetic, morphological, biogeographic, ecological, and behavioral evidence indicates that reclassifying Bwindi gorillas as taxonomically distinct from those in the Virungas is not well supported and needs further study. Because taxonomy provides the basis of conservation management policies, a cautious and conservative approach to the subspecies question is warranted in the case of endangered primate populations.     

Mapping of <Emphasis Type="Italic">taiep</Emphasis> rat phenotype to rat Chromosome 9

The taiep mutant rat was first described in a colony of Sprague-Dawley rats at the University of Puebla in 1989, with an autosomal recessive inherited pattern. taiep is an acronym for the progressive neurologic deficits that the rat develops, i.e., t = trembling (3–4 weeks), a = ataxia (at 4 months), i = immobility (5–6 months), e = epilepsy (5–6 months), and p = paresis (7 months onwards). Thus, mutant rats are first identified by a tremor at 3–4 weeks of age that is followed by a progressive neurological worsening (Holmgren et al. 1989; Lunn et al. 1997). The cause of the neurological symptoms is an early failure of normal myelination of the central nervous system (CNS) followed by progressive demyelination of certain CNS tracts (Lunn et al. 1997). We have been exploring the underlying pathophysiology of the mutant and have determined that the myelin defect results from the progressive accumulation of microtubules in oligodendrocytes, the myelin-producing cells of the CNS (Song et al. 1999). Microtubules are the major component of the cytoskeleton of this and many other cells of the body, and microtubule-based transport of protein and mRNA is essential for normal cell function. There is no direct human counterpart of the taiep rat. Nonetheless, providing an understanding of the control of microtubule dynamics in the oligodendrocyte will be highly relevant to our knowledge of the cell biology of the myelinating cell of the CNS. This information is of great relevance to the function of the cell in human myelin disorders and in experimental remyelination. As the taiep rat apparently has a primary disorder in the oligodendrocyte cytoskeleton, it is an ideal model in which to study this process. This information may also be a key to the complete understanding of the mechanism of microtubule assembly/disassembly in many cell types.     

Location and nucleotide sequence of a tobacco chlorophlast DNA segment capable of replication in yeast

Summary A 1.7 Kbp EcoRI fragment of Nicotiana tabacum chloroplast DNA cloned in YIp5, consisting of pBR322 and the yeast ura3 gene, possessed ars (autonomously replicating sequences) activity in Saccharomyces cerevisiae. This fragment was located in the small single copy region proximal to the 23S rRNA gene.Sequences responsible for potential ars activity were narrowed to about 350 base pairs, where clusters of nucleotides similar to a consensus sequence (11 bp) essential for several yeast ars (Broach et al. 1982), to the stem-and-loop structure typical of yeast ars3 (Feldmann et al. 1981), and regions surrounding the replication origin of mitochondrial DNAs of HeLa Cells (Crews et al. 1979) and yeast (de Zamaroczy et al. 1981) can be observed.Abbreviations ctDNA chloroplast DNA - Kbp kilobase pairs     

茶树品种(品系)芽叶色泽表型遗传数量分类研究

为探讨茶树春梢芽叶色泽的数量遗传特性,应用色差仪测定68份茶树品种(品系)春梢芽叶的色泽表型值并进行数量分类研究。结果表明,对叶色差异度较高的第1叶位L~*、a~*、b~*、C、H值进行分层聚类分析,可将芽叶色泽划分为紫色系、黄色系和绿色系等7个色系。色系分类与CIELab颜色体系的对应性较强,色系划分符合茶树品种(品系)叶色表型特点。茶树品系的叶色表型较丰富,叶色在L~*、a~*、b~*值三维坐标空间整体呈带域分布,色系由紫色系、紫红色系、紫绿色系向黄色系、黄绿色系、绿色系变化。春梢芽叶色泽表型数据的遗传数量分类与色系判定为茶树特异芽叶色泽育种提供科学依据。     

An improved method for large-scale purification of recombinant human glucagon

Glucagon was expressed inEscherichia coli as a fusion protein including the glucagon sequence [Ishizakiet al. (1992),Appl. Microbiol. Biotechnol.36, 483–486]. The high-level expression of a protein inE. coli often results in an insoluble aggregate called an inclusion body containing a fusion protein. In our previous report [Yoshikawaet al. (1992),J. Protein Chem. 11, 517–525], we solubilized this inclusion body by using guanidinium chloride. However, the existence of denaturant caused problems such as a low proteolytic activity for transforming the fusion protein into glucagon and complicated purification methods. We tried to improve the method to enable large-scale purification. At alkaline pH, the inclusion body could be solubilized to a high concentration and cleaved by amino acid-specific endopeptidases. By utilizing isoelectric precipitations as a new economical purification method for glucagon from intermediates, the glucagon obtained was shown to be over 99.5% pure by analytical RP-HPLC. The yield was almost equal that of our previous method, and the glucagon produced was chemically and biochemically equivalent to natural glucagon.     

Problems in the experimental determination of substrate-specific H+/O ratios during respiration

Krabet al. (1984) have recently tried to resolve the long-standing controversy as to whether the mechanistic H⁺/O coupling ratio for electrons passing through sites II and III of the mammalian electron transport chain to O₂ is 6 or 8. Using a mathematical model they concluded that the higher number reported by Costaet al. (1984) was an overestimate because of the unaccounted for delayed response of the O₂ electrode. Responding to criticisms of Lehningeret al. (1985), they have recently used (Krab and Wikström, 1986) an improved mathematical model which shows that the higher number found by Costaet al. was probably due to an inadequate accounting for the effects of the proton leak process which accompanies the translocation process. The impression is left that the situation is now resolved in favor of the lower number. We agree that the procedures of Costaet al. do not properly account for the leak process, and provide further evidence in this paper of the magnitude of the problem. However, we disagree that the number 6.0, favored by Wikströmet al., rests on any more solid experimental support. We provide evidence here for this conclusion and raise the question as to whether or not any unique, fixed, integral number exists for the H⁺/O ratio accompanying the oxidation of a particular substrate.