Exchanges across land-water-scape boundaries in urban systems: strategies for reducing nitrate pollution.

Conservation in urban areas typically focuses on biodiversity and large green spaces. However, opportunities exist throughout urban areas to enhance ecological functions. An important function of urban landscapes is retaining nitrogen thereby reducing nitrate pollution to streams and coastal waters. Control of nonpoint nitrate pollution in urban areas was originally based on the documented importance of riparian zones in agricultural and forested ecosystems. The watershed and boundary frameworks have been used to guide stream research and a riparian conservation strategy to reduce nitrate pollution in urban streams. But is stream restoration and riparian-zone conservation enough? Data from the Baltimore Ecosystem Study and other urban stream research indicate that urban riparian zones do not necessarily prevent nitrate from entering, nor remove nitrate from, streams. Based on this insight, policy makers in Baltimore extended the conservation strategy throughout larger watersheds, attempting to restore functions that no longer took place in riparian boundaries. Two urban revitalization projects are presented as examples aimed at reducing nitrate pollution to stormwater, streams, and the Chesapeake Bay. An adaptive cycle of ecological urban design synthesizes the insights from the watershed and boundary frameworks, from new data, and from the conservation concerns of agencies and local communities. This urban example of conservation based on ameliorating nitrate water pollution extends the initial watershed-boundary approach along three dimensions: 1) from riparian to urban land-water-scapes; 2) from discrete engineering solutions to ecological design approaches; and 3) from structural solutions to inclusion of individual, household, and institutional behavior.

Replication protein A is sequentially phosphorylated during meiosis.

Phosphorylation of the cellular single-stranded DNA-binding protein, replication protein A (RPA), occurs during normal mitotic cell cycle progression and also in response to genotoxic stress. In budding yeast, these reactions require the ATM homolog Mec1, a central regulator of the DNA replication and DNA damage checkpoint responses. We now demonstrate that the middle subunit of yeast RPA (Rfa2) becomes phosphorylated in two discrete steps during meiosis. Primary Rfa2 phosphorylation occurs early in meiotic progression and is independent of DNA replication, recombination and Mec1. In contrast, secondary Rfa2 phosphorylation is activated upon initiation of recombination and requires Mec1. While the primary Rfa2 phosphoisomer is detectable throughout most of meiosis, the secondary Rfa2 phosphoisomer is only transiently generated and begins to disappear soon after recombination is complete. Extensive secondary Rfa2 phosphorylation is observed in a recombination mutant defective for the pachytene checkpoint, indicating that Mec1-dependent Rfa2 phosphorylation does not function to maintain meiotic delay in response to DNA double-strand breaks. Our results suggest that Mec1-dependent RPA phosphorylation could be involved in regulating recombination rather than cell cycle or meiotic progression.

Components of length growth variation in infants from the same population but different environments.

Growth in length of infants in Khartoum is profoundly related to whether or not their mothers are in paid employment. Some of this relationship appears to be due to levels of infectious disease, which are much higher in the infants of working mothers. Exponential decay curves do not fit longitudinal measurements of length growth particularly well in either group of infants, but they do yield distinctive temporal patterns of residuals. Simulation demonstrates that these patterns accord with a "catch-down" form of growth in infants of working mothers and a "catch-up" form in infants of housewives. Attempts to allow for these phenomena by providing for the opportunity of a single pulse of negative sine form in the former and a similar pulse of positive sine form in the latter reduces the level of residuals and tends to reduce temporal pattern in them. Most strikingly, it leads to average growth curves for the two groups of children becoming almost identical. It is hypothesized that these decay curves represent the underlying curves of infant growth in Khartoum free from the effects of environmental variation.

Growth of specific muscle strength between 6 and 18 years in contrasting socioeconomic conditions.

The influence of sex, age, and socioeconomic conditions on specific grip strength of 6-18-year-old individuals was studied among 1,704 males and 1,956 females belonging to the so-called "Cape Coloured" community in the western part of South Africa. Half of the participants of both sexes came from communities in the Greater Cape Town area where living conditions are comparable to those of middle-class First World communities (high SES). The other half came from the poorest rural communities of Klein Karoo (low SES). Arm circumferences, triceps skinfold thickness, and grip strength of the right and of the left hand were greater in individuals from high SES at all ages. Females within each SES group had skinfolds thicker than males, especially at older ages, and were weaker. Specific grip strength (SS), estimated as grip strength per unit area of cross section of the fat-free arm, increased with age in each group, was greater in males, and was significantly lower in low SES groups, than in the high SES ones, especially during and after puberty. It seems that SES difference in SS will persist into adulthood. Sexual differences in SS can be attributed to hormonal differences; while the SS increase with age and the difference between SES groups find no clear explanation in current theories of muscle growth and development. Since the speed of neuromuscular reaction observed in our participants is slower among low SES individuals, it seems that the difference in neuromuscular control of strength may be responsible for our findings. Differences in muscle metabolism and hormonal regulation must also be considered.

Phosphorylation of the replication protein A large subunit in the Saccharomyces cerevisiae checkpoint response.

The checkpoint mechanisms that delay cell cycle progression in response to DNA damage or inhibition of DNA replication are necessary for maintenance of genetic stability in eukaryotic cells. Potential targets of checkpoint-mediated regulation include proteins directly involved in DNA metabolism, such as the cellular single-stranded DNA (ssDNA) binding protein, replication protein A (RPA). Studies in Saccharomyces cerevisiae have revealed that the RPA large subunit (Rfa1p) is involved in the G1 and S phase DNA damage checkpoints. We now demonstrate that Rfa1p is phosphorylated in response to various forms of genotoxic stress, including radiation and hydroxyurea exposure, and further show that phosphorylation of Rfa1p is dependent on the central checkpoint regulator Mec1p. Analysis of the requirement for other checkpoint genes indicates that different mechanisms mediate radiation- and hydroxyurea-induced Rfa1p phosphorylation despite the common requirement for functional Mec1p. In addition, experiments with mutants defective in the Cdc13p telomere-binding protein indicate that ssDNA formation is an important signal for Rfa1p phosphorylation. Because Rfa1p contains the major ssDNA binding activity of the RPA heterotrimer and is required for DNA replication, repair and recombination, it is possible that phosphorylation of this subunit is directly involved in modulating RPA activity during the checkpoint response.

Identification and characterization of a single-stranded DNA-binding protein from the archaeon Methanococcus jannaschii.

Single-stranded DNA-binding proteins (SSBs) play essential roles in DNA replication, recombination, and repair in bacteria and eukarya. We report here the identification and characterization of the SSB of an archaeon, Methanococcus jannaschii. The M. jannaschii SSB (mjaSSB) has significant amino acid sequence similarity to the eukaryotic SSB, replication protein A (RPA), and contains four tandem repeats of the core single-stranded DNA (ssDNA) binding domain originally defined by structural studies of RPA. Homologous SSBs are encoded by the genomes of other archaeal species, including Methanobacterium thermoautotrophicum and Archaeoglobus fulgidus. The purified mjaSSB binds to ssDNA with high affinity and selectivity. The apparent association constant for binding to ssDNA is similar to that of RPA under comparable experimental conditions, and the affinity for ssDNA exceeds that for double-stranded DNA by at least two orders of magnitude. The binding site size for mjaSSB is approximately 20 nucleotides. Given that RPA is related to mjaSSB at the sequence level and to Escherichia coli SSB at the structural level, we conclude that the SSBs of archaea, eukarya, and bacteria share a common core ssDNA-binding domain. This ssDNA-binding domain was presumably present in the common ancestor to all three major branches of life.

The small child: anthropometric and physical performance characteristics of short-for-age children growing in good and in poor socio-economic conditions.

OBJECTIVE: To ascertain whether individuals whose short stature is caused by poor living conditions differ in their other biological characteristics from those individuals who grow short, supposedly due to their genetic endowment, under favourable environmental conditions. DESIGN: From the growth survey of nearly 4000 children of the 'Cape Coloured' community aged 6-18 y half of whom lived in the middle-class suburbs of Cape Town [high socio-economic status (SES)], the other half who lived in poor rural settlements (low SES), boys and girls of similarly short stature were selected. Average body height by sex and age in each selected sample was close to the 5th percentile of height distribution among all high SES individuals surveyed. SETTING: Cape Town and neighbouring rural settlements in South Africa. SUBJECTS: Four thousand children of the Cape Coloured community aged 6-18 y. Among these were 109 high SES boys, 123 high SES girls, 302 low SES boys and 344 low SES girls selected for short stature study. INTERVENTIONS: Anthropometric measurements in public schools with appropriate permissions. RESULTS: Both short stature males and females from the low SES group tended to be significantly (P < 0.05) different from the high SES group of comparable stature in having lower body weights, shorter limbs, longer trunks, narrower shoulders, hips and chests, thinner skinfolds, smaller arm circumferences, and were very notably weaker and had poorer neuromuscular reaction times. Biepicondylar widths and pulse rates were little different between groups. Genetically short children do differ in various anthropometric and functional traits from those who are small from environmental causes.

Effects of early disease on later growth, and early growth on later disease, in Khartoum infants.

Longitudinal data of monthly length and weight measurements, and fortnightly infectious disease records of 167 infants in Khartoum townships, have been examined to determine: (1) whether disease experience in the first 6 months of life is related to growth in the second 6 months and (2) whether growth in the first period is related to disease experience in the second. Using regression techniques to allow for disease measure correlations in the two periods and other environmental factors that are associated with growth and disease, it is concluded that both length and especially weight growth are affected adversely by previous illness. On the other hand there is almost no evidence that early growth is associated with later disease susceptibility.

The ATM homologue MEC1 is required for phosphorylation of replication protein A in yeast.

Replication protein A (RPA) is a highly conserved single-stranded DNA-binding protein, required for cellular DNA replication, repair, and recombination. In human cells, RPA is phosphorylated during the S and G2 phases of the cell cycle and also in response to ionizing or ultraviolet radiation. Saccharomyces cerevisiae exhibits a similar pattern of cell cycle-regulated RPA phosphorylation, and our studies indicate that the radiation-induced reactions occur in yeast as well. We have examined yeast RPA phosphorylation during the normal cell cycle and in response to environmental insult, and have demonstrated that the checkpoint gene MEC1 is required for the reaction under all conditions tested. Through examination of several checkpoint mutants, we have placed RPA phosphorylation in a novel pathway of the DNA damage response. MEC1 is similar in sequence to human ATM, the gene mutated in patients with ataxia-telangiectasia (A-T). A-T cells are deficient in multiple checkpoint pathways and are hypersensitive to killing by ionizing radiation. Because A-T cells exhibit a delay in ionizing radiation-induced RPA phosphorylation, our results indicate a functional similarity between MEC1 and ATM, and suggest that RPA phosphorylation is involved in a conserved eukaryotic DNA damage-response pathway defective in A-T.

Crystal structure of bacteriophage T4 deoxynucleotide kinase with its substrates dGMP and ATP.

NMP kinases catalyse the phosphorylation of the canonical nucleotides to the corresponding diphosphates using ATP as a phosphate donor. Bacteriophage T4 deoxynucleotide kinase (DNK) is the only member of this family of enzymes that recognizes three structurally dissimilar nucleotides: dGMP, dTMP and 5-hydroxymethyl-dCMP while excluding dCMP and dAMP. The crystal structure of DNK with its substrate dGMP has been determined at 2.0 A resolution by single isomorphous replacement. The structure of the ternary complex with dGMP and ATP has been determined at 2.2 A resolution. The polypeptide chain of DNK is folded into two domains of equal size, one of which resembles the mononucleotide binding motif with the glycine-rich P-loop. The second domain, consisting of five alpha-helices, forms the NMP binding pocket. A hinge connection between the domains allows for large movements upon substrate binding which are not restricted by dimerization of the enzyme. The mechanism of active centre formation via domain closure is described. Comparison with other P-loop-containing proteins indicates an induced-fit mode of NTP binding. Protein-substrate interactions observed at the NMP and NTP sites provide the basis for understanding the principles of nucleotide discrimination.

Crystallization and preliminary X-ray analysis of bacteriophage T4 deoxynucleotide kinase.

T4 deoxynucleotide kinase catalyzes the phosphorylation of 5-hydroxymethyldeoxycytidylate, dTMP and dGMP while excluding dCMP and dAMP. In order to understand the mechanism of this remarkable specificity, the enzyme was over-expressed in Escherichia coli, purified and crystallized for X-ray diffraction analysis. The crystals belong to the monoclinic space group C2 with cell dimensions a = 155.2, b = 58.5, c = 75.7 A, beta = 108.1 degrees. There are two protein monomers in the asymmetric unit related by a twofold axis. Diffraction data to 2.0 A resolution have been collected.

The DNA-activated protein kinase is required for the phosphorylation of replication protein A during simian virus 40 DNA replication.

The 32-kDa subunit of replication protein A (RPA) is phosphorylated during the S phase of the cell cycle in vivo and during simian virus 40 DNA replication in vitro. To explore the functional significance of this modification, we purified a HeLa cell protein kinase that phosphorylates RPA in the presence of single-stranded DNA. By several criteria we identified the purified enzyme as a form of the DNA-activated protein kinase (DNA-PK), a previously described high molecular weight protein kinase that is capable of phosphorylating a number of nuclear DNA binding proteins. Phosphorylation of RPA by DNA-PK is stimulated by natural single-stranded DNAs but not by homopolymers lacking secondary structure. Studies with the simian virus 40 model system indicate that DNA-PK is required for DNA-replication-dependent RPA phosphorylation. Depletion of the kinase activity, however, has no effect on the extent of DNA replication in vitro. Our data support a model in which phosphorylation of RPA by DNA-PK is activated by formation of replication intermediates containing single- and double-stranded regions. This event may be involved in a signaling mechanism that coordinates DNA replication with the cell cycle.

Comparative length and weight growth in Khartoum infants.

First-year postnatal growth of supine length and body weight of a group of Khartoum infants is analysed in terms of the correlation coefficients between birth dimension on the one hand and (1) subsequent distance growth at monthly intervals and (2) the 11-monthly increments on the other. It is shown that the whole of the first-year length growth is dominated by catch-up/catch-down processes which must equally affect the infants. Weight growth is similarly affected, but to a lesser degree and for a shorter time. After 6 months of age some of the influences that determined prenatal weight growth reappear. Using data extracted from fitting growth curves to each individual the correlational analysis suggests a growth pulse in both length and weight, which is responsible for distance catch-up.

A path analysis of some determinants of infant growth in Khartoum.

The interrelationships between some socioeconomic and behavioural characteristics of mothers and the growth and illness experience of their infants in poor Khartoum townships have been examined by path analysis. For both infant body weight and supine length it is shown that weaning age and illness experience are important determinants of growth. It is also shown that greater maternal income has disadvantageous effects through encouraging early weaning. Most importantly the infants of housewives have later weaning, less illness and greater weight and length at 1 year of age than the infants of mothers with jobs, showing the value of time for caring in the prevailing environmental circumstances.

PIP: Path analysis was performed for infant weight and length among 96 mother-infant pairs in the study sample. The aim was to uncover causal connections between maternal income, maternal occupation, birth weight, weight at 2 months, weaning age, experience of illness from 3 to 12 months of age, and weight at 1 year. The sample was comprised of infants and mothers who were 18-30 years old, in the third trimester of pregnancy, and with 0-3 prior births. The results revealed little difference between the observed correlations and those predicted by the models. The weight model showed the expected strong positive path between birth weight and weight at 2 months and the negative path between illness experience and weight at 1 year. Late weaning appears to protect from illness and contributes to first year growth independently. Heavy infants tend to be weaned late. Heavy infants also have less illness independent of early weight gain. Infants of homemakers had higher birth weights, lower 2-month birth weights, greater 1-year birth weights, and less illness than the infants of paid working women. Homemaker status also affected weaning age. In the length model, illness affected length at 1 year, and weaning and length were not directly connected. Homemakers' infants were shorter at birth and at 2 months, but longer at 1 year than the infants of paid working women. The weight model was better at characterizing pathways and had better goodness of fit (x2 = 9.08, p 0.05). The greater health of at-home mothers in poor economic settings has important policy implications.

Motherhood and infant health in Khartoum.

The results of an analysis of the growth and illness experience of a group of infants in Khartoum townships illustrate the remarkable benefits of being the infant of a "housewife" rather than the infant of a mother who works. These benefits occurred despite the poorer domestic environments of the housewives.

PIP: Path analysis of growth and illness experiences among study infants reveals that housewives' infants have substantially less illness and better first year increases in weight and length than infants of working mothers in a poor township in Khartoum, Sudan. The analysis is based on a sample of 120 mother-infant pairs and information on monthly illness experiences in two week periods throughout the first year of life. The sample includes 36 housewives and 84 working mothers. The mean illness duration is reported to be an increase to 4 days per month at 7 months for infants of working mothers. Illness duration thereafter remains at over 3 days per month. Differences in weaning time are statistically significant. Infants are weaned between 156.8 +or- 61.5 days for infants of housewives compared to 133.34 +or- 69.7 days for infants of working mothers. Few differences in body weight and supine length occur during the early growth period. After 6 months, growth curves diverge, such that by 12 months there is a statistically significant difference of 2.1 kg between infants. Infants of working mothers are initially longer, converge with housewives' infants by 5-6 months, and are shorter by 12 months by 3.8 cm. Weight and length, except length at 11-12 months, varies less at all ages among infants of housewives. Initial differences in length are attributed to gestation age. The evidence is considered supportive of a health advantage among infants of housewives. The study would be improved with knowledge about the health of mothers. Information on maternal income indicates that housewives have less income. The path analysis finds the "housewife" effect on infant health independent of income. Housewives are found to have more adverse living conditions (poorer housing, sanitation, hygiene, and water supply). The evidence suggests the importance of maternal behavior.

Chemical modification of bacteriophage T4 deoxynucleotide kinase. Evidence of a single catalytic region.

The mechanism underlying the unusual specificity of bacteriophage T4 deoxynucleotide kinase, which catalyzes the phosphorylation of 5-hydroxymethyldeoxycytidylate, dTMP, and dGMP, has been investigated by chemical modification of the protein. Pyridoxal 5'-phosphate inactivates deoxynucleotide kinase by modifying a single lysine out of the 17 per monomer. Lysine 10 has been tentatively identified as the site of modification, although the possibility of mutually exclusive reactive residues has not been eliminated. Diethylpyrocarbonate also inactivates the enzyme, suggesting that histidine plays a role in catalytic function. With either reagent, the three activities are lost at equal rates, supporting the contention that one active site is responsible for the exclusive phosphorylation of three dissimilar deoxynucleotides. These studies also identify two distant regions of the primary sequence that are likely to be closely associated in the active region of the folded protein.

Interrelations between growth, weaning and disease experience in Khartoum infants.

The monthly measurements of body weight and supine length from birth to 1 year of 203 infants in Khartoum were analysed to see whether growth characteristics affected the timing of weaning and the amount of illness experience. The study clearly shows that neonates who are long for their age tend to be weaned early, and this causes them to experience a greater average monthly 'illness experience' and slower subsequent growth. However, a multivariate analysis indicates that it is correlated environmental variables rather than length growth itself which primarily determines weaning age. Weight growth does not show these relationships but there is strong evidence that late weaning, independently of disease experience, promotes post-natal weight growth.