Genetic studies of low-abundance human plasma proteins. XIII. Population genetics of C1R complement subcomponent and description of new variants

Isoelectric focusing and immunoblotting reveals considerable biochemical and genetic variation in the C1R subcomponent of the first complement component. The nature of the intraindividual biochemical variation can be explained by differences in sialic acid content because after digestion with neuraminidase the terminal sialic acids are removed to yield a single major band corresponding to the C1R polypeptide. Plasma samples from a large number of different ethnic groups, consisting of U.S. whites, U.S. blacks, Nigerian blacks, and Inuit, Aleut, and Amerindian populations from the Western Hemisphere have revealed genetically determined charge variation with heterozygous phenotypes consisting of two major asialo bands, indicating that the underlying variation is not due to variation in sialic acid content. Two previously reported common alleles, C1R*1 and CIR*2, have been observed in all studied populations, the notable exception being the Dogrib Indian population, which is devoid of the C1R*2 allele. Several new alleles--designated C1R*3, C1R*4, C1R*5, C1R*6, and C1R*7-have been observed, with variable frequencies ranging from the occurrence in a single individual and related family members to the polymorphic occurrence of certain alleles in several populations. Of these new alleles, the C1R*5 is of considerable interest in population and anthropological genetics studies. The C1R*5 allele is widely distributed, at a frequency of .03 to .17, in all of the North American aboriginal populations screened. This allele is not present in U.S. whites but is present at a polymorphic frequency in U.S. and Nigerian blacks.(ABSTRACT TRUNCATED AT 250 WORDS)     

Genetic studies of human apolipoproteins. IX. Apolipoprotein D polymorphism and its relation to serum lipoprotein lipid levels.

Apolipoprotein D (APO D) is a constituent of plasma high-density lipoproteins. Its precise role in lipid metabolism is not well established, though it may be involved in cholesterol esterification and cholester ester transport to the liver for catabolism. No genetic polymorphism has been reported in the APO D gene product. To investigate the extent of genetic variation at the APO D structural locus, we have developed an isoelectric focusing-immunoblotting technique and have screened a large number of plasma samples from U.S. whites, U.S. blacks, Nigerian blacks, the Aleuts of the Pribilof Islands, Eskimo groups from Kodiak Island and St. Lawrence Island, and Amerindian populations from Mexico and Canada. Except for the U.S. blacks and Nigerian blacks, the APO D locus is monomorphic in all other population groups tested. In populations with black ancestry, the products of two alleles, APO D*1 and APO D*2, have been observed at respective allele frequencies .987 and .013 in U.S. blacks and .978 and .022 in Nigerian blacks. The detection of a unique protein polymorphism in blacks makes APO D a useful black marker of significance in anthropogenetics and racial admixture studies. In addition to the interindividual variation observed, APO D reveals extensive intraindividual molecular variation with a multiple banding pattern. The basis of this molecular variation is explained, in part, by variation in the number of terminal sialic acid residues. We have investigated the effect of the APO D polymorphism on triglycerides, total cholesterol, LDL-, VLDL-, HDL-, and HDL3 cholesterol in 352 Nigerian blacks (190 males and 162 females).(ABSTRACT TRUNCATED AT 250 WORDS)     

Genetic studies of human apolipoproteins. I. Polymorphism of apolipoprotein A-IV.

Genetic polymorphism of human apolipoprotein A-IV has been detected by means of a simple and rapid one-dimensional isoelectric-focusing technique followed by immunoblotting. In plasma samples of normal U.S. whites and blacks, the specificity and sensitivity of the technique have been demonstrated to elucidate biochemical and genetic variation present in the APO A-IV molecule. Two common alleles, APO A-IV 1 and APO A-IV 2, have been observed with respective frequencies of .909 and .088 in whites and .961 and .035 in blacks. In addition, the products of two rare alleles designated APO A-IV 3 and APO A-IV 4 also have been observed. Family studies show autosomal codominant transmission of four alleles coded by a single structural locus.     

Genetic studies of human apolipoproteins. IV. Structural heterogeneity of apolipoprotein H (beta 2-glycoprotein I).

Human beta 2-glycoprotein I has recently been identified as a component of several human plasma lipoprotein fractions and therefore termed as apolipoprotein H. Its metabolic function in lipid metabolism is not known with certainty, though it may be involved in very-low-density-lipoprotein metabolism. Previously, inherited quantitative variation in beta 2-glycoprotein I has been suggested in man. In this investigation, we document the evidence of genetically determined structural polymorphism of apolipoprotein H or beta 2-glycoprotein I by using thin-layer polyacrylamide isoelectric focusing gels followed by immunological identification by double antibody staining. The apolipoprotein H structural locus is characterized by the occurrence of three common alleles in U.S. whites and blacks. The frequency distributions of the three alleles designated APO H1, APO H2, and APO H3 are .059, .882, and .059 in whites and .017, .902, and .068 in blacks, respectively. In addition, the gene product of a fourth allele, APO H4, has been observed at polymorphic frequency in black individuals and may represent a black marker variant. Family data confirm the hypothesis of four alleles at a single APO H gene locus with an autosomal codominant pattern of inheritance.     

Genetic studies of low abundance human plasma proteins. II. Population genetics of coagulation factor XIIIB.

Plasma samples from a large number of different ethnic groups, consisting of U.S. whites, U.S. blacks, Eskimos from Kodiak and St. Lawrence Island, Aleuts of the Pribilof Islands, and three Amerindian groups from Canada and Mexico have been analyzed by isoelectric focusing followed by immunoblotting to determine the magnitude of genetic variation at the F XIIIB structural locus. The synthesis of published data and our new data demonstrate remarkable variation in the distribution of the three common alleles at this locus and establishes F XIIIB as an extremely informative marker for population differentiation and evolutionary studies. Genetic distance analysis based on this variation separates Caucasian, black, and Mongoloid populations into three distinct clusters.     

Comparability of Resting Energy Expenditure in Nigerians and U.S. Blacks

Objective: To determine the influence of environmental factors on resting energy expenditure (REE) and its relationship to adiposity in two populations of West African origin, Nigerians and U.S. blacks. Research Methods and Procedures: REE and body composition were measured in a cross‐sectional sample of 89 Nigerian adults (39 women and 50 men), and 181 U.S. black adults (117 women and 65 men). Both groups represent randomly selected population samples. REE was measured by indirect calorimetry after an overnight fast in both sites using the same instrument. Body composition was estimated using bioelectrical impedance analysis (BIA) in 72 Nigerians and 156 U.S. participants. Multivariate regression analysis was used to determine the significant predictors of REE. The analyses were repeated in a set of 17 Nigerians and 28 U.S. blacks in whom body composition was measured using deuterium dilution. Results: U.S. black adults were significantly heavier and had both more fat‐free mass (FFM) and body fat than Nigerians. FFM was the only significant determinant of REE in both population groups, whether body composition was measured using BIA or deuterium dilution. The relationship between REE and body composition did not differ by site. There was no relationship between REE and adiposity. Discussion: Differences in current environmental settings did not impact REE. The differences observed in mean levels of body fat between Nigerians and U.S. blacks were not the result of differences in REE adjusted for body composition.     

Caucasian genes in American blacks: new data.

Data on 15 polymorphic protein-coding loci are used to estimate the proportion of Caucasian genes in U.S. blacks from the greater-metropolitan area of Pittsburgh. Allele frequencies from U.S. whites of the same region and from a sample of Nigerians are used as representatives of the genetic contributions of the source populations between which admixture has occurred. These materials provide 18 unique variants that occur exclusively in the blacks and 5 variants that are restricted to the Caucasians only. As a result, when all segregating alleles (52) at these 15 loci are considered, the proportion (mean +/- SE) of Caucasian genes in U.S. blacks (25.2% +/- 2.7%) is estimated with a precision much better than that of all other previous estimates. The estimate based on the frequencies of these 18 unique variants of African origin (24.8% +/- 6.2%) is also consistent with the pooled estimate obtained from the 15 loci by the weighted least-square method. The homogeneity of locus-specific estimates of admixture indicates that these loci are appropriate for studying the proportion of black genes in any admixed population involving African admixture. The advantages of employing such loci for genetic-epidemiologic studies in U.S. blacks is discussed in the context of the availability of these large number of unique African variants at these protein loci.     

Genetic studies of human apolipoproteins. XX. Genetic polymorphism of apolipoprotein J and its impact on quantitative lipid traits in normolipidemic subjects.

Apolipoprotein J (apo J) is a newly identified member of a growing family of proteins associated with various lipoprotein particles. Apo J is a glycoprotein which exists in the plasma associated with high-density lipoprotein subfractions which also contain apo A-I and cholesteryl ester transfer protein (CETP). We have investigated the possible existence of genetic polymorphism at the apo J structural locus and have evaluated its role in lipid metabolism. By employing isoelectric focusing and immunoblotting techniques, we have screened plasma or serum samples from six population groups: U.S. whites, Amerindians, Eskimos, New Guineans, U.S. blacks, and Nigerian blacks. Apo J revealed a common two-allele polymorphism only in populations with African ancestry and was found to be monomorphic in all other population groups tested. The genetic basis of the two alleles designated--APO J*1 and APO J*2, at a single structural locus, apo J-- was confirmed in a large number of segregating families. In the U.S. blacks, the frequencies of the APO J*1 and APO J*2 alleles were .76 and .24, respectively, and in the Nigerian blacks these values were .72 and .28, respectively. In addition, a single example of a rare allele designated APO J*3 was also encountered in the U.S. black sample. In Nigerian blacks, the apo J polymorphism's impact on seven quantitative lipid traits--total cholesterol, LDL-cholesterol, HDL-cholesterol, HDL3-cholesterol, HDL2-cholesterol, VLDL-cholesterol, and triglycerides--was investigated. No significant impact of the apo J polymorphism was observed for any of these lipid traits.     

An Analysis of the Proportion of the U.S. Population that Ingests Soil or Other Non-Food Substances

Reliable quantitative data are lacking that document the prevalence of ingestion of soil and other “non-food” substances among U.S. children and adults. This article explores the proportion of the U.S. population that ingests substances such as soil, clay, starch, paint, or plaster. We compiled data from the National Health and Nutrition Examination Survey (NHANES) collected from years 1971–1975 (NHANES I) and 1976–1980 (NHANES II) because these particular surveys asked participants specific questions about non-food ingestion practices. We examined the prevalence of the behavior across multiple demographic variables, such as age, gender, education, and income level. Approximately 1% (NHANES II) to 2.5% (NHANES I) of the U.S. population ingests some type of non-food substance. The most notable variation across the demographic subgroups studied was the difference in estimated prevalence among young children (1 to <3 years) compared to older children and adults. Estimated prevalence was also higher among blacks compared to whites and within lower compared to higher socioeconomic groups. This analysis helps fill data gaps on the relative pattern of non-food ingestion practices on a national scale. This information provides perspective for risk assessors when evaluating exposure variables and for risk managers when weighing risk management alternatives.     

Further studies on the genetic control of chicken plasma alkaline phosphatase isozyme.

The effects of neuraminidase treatment on the electrophoretic pattern of alkaline phosphatase (AP) isozymes and AP activity were investigated in chicken plasma. AP comprised three isozymes. The zymogram of an individual chicken plasma had two bands, either the faster (F) or the slower (S) moving band by isozyme types and the B band irrespective of isozyme types. Mobility of the S band and AP activity in chicken plasma were not affected by neuraminidase treatment. The treatment has a reduced migration rate of the F band equal to that of the S band and the B band of both types closer to the origin. The genetic control of these bands is discussed.     

Alpha1-antitrypsin: further genetic heterogeneity revealed by isoelectric focusing.

Alpha1-antitrypsin is a major human serum protein that shows an extensive polymorphism. Genetic heterogeneity has previously been demonstrated by starch gel electrophoresis. By applying analytical isoelectric focusing (pH 3.5--5.0) to this system, we found a common variant, Pi M3, with an isoelectric point between those of Pi M1 and Pi M2. The gene frequency of this variant was .11 in U.S. whites and .054 in blacks. When PiM3 and PiM1 are included in the Pi system, the heterozygosity at the Pi locus is five times greater in whites and 10 times greater in blacks than that detected by earlier electrophoretic techniques.     

Characterization of two gangliosides from human leukemic polymorphonuclear leukocytes.

Eight bands of gangliosides, from human polymorphonuclear leukocytes were demonstrated by thin-layer chromatography. Bands 4 and 5 were isolated and purified in sufficient amounts to allow their biochemical identification by thin-layer chromatography, gas chromatography and sequential action of glycosidases and neuraminidase. The major ganglioside was characterised as N-acetylneuraminylgalactosyl-beta-N-acetylglucosaminyl-beta-galactosyl-beta-glucosylceramide. A second ganglioside was tentatively identified as N-acetylneuraminyl-galactosyl-beta-N-acetylglucosaminyl-beta-(N-acetylneuraminyl)galactosyl-beta-glucosylceramide. Both gangliosides isolated were hydrolysed by neuraminidase. However, treatment of the intact cells with neuraminidase did not alter the ganglioside pattern.     

Seasonal wing colour plasticity varies dramatically between buckeye butterfly populations in different climatic zones

1. In the present study, how seasonal wing colour plasticity in buckeye butterflies (Junonia coenia Hübner) differs between populations in different climatic regions, and whether these differences are explained by regional environmental differences or reflect genetic divergences in plasticity is addressed. 2. Using museum specimen data, it is shown that buckeye wing colour variation is much greater in populations from the humid and subtropical climates of the eastern U.S.A. than in populations from the desert and mediterranean climates of the western U.S.A. 3. Museum specimen data further show that wing colour in eastern populations is strongly correlated with seasonal variation in day length and temperature, whereas wing colour in western populations is only weakly associated with these cues. 4. Controlled incubator experiments comparing Southern California and North Carolina populations suggest that regional differences in wing pattern variation are attributable to dramatically increased robustness to environmental variation in western populations. 5. Together these experiments show that while phenotypic variation can be influenced by environmental cues, the range of this variation can genetically diverge between populations in different regions.     

Genetic population structure of spotted seatrout Cynoscion nebulosus along the south‐eastern U.S.A.

Analyses of the genetic population structure of spotted seatrout Cynoscion nebulosus along the south‐eastern U.S. coast using 13 microsatellites suggest significant population differentiation between fish in North Carolina (NC) compared with South Carolina (SC) and Georgia (GA), with New River, NC, serving as an area of integration between northern and southern C. nebulosus. Although there is a significant break in gene flow between these areas, the overall pattern throughout the sampling range represents a gradient in genetic diversification with the degree of geographic separation. Latitudinal distance and estuarine density appear to be main drivers in the genetic differentiation of C. nebulosus along the south‐eastern U.S. coast. The isolation‐by‐distance gene‐flow pattern creates fine‐scale differences in the genetic composition of proximal estuaries and dictates that stocking must be confined to within 100 km of the location of broodstock collection in order to maintain the natural gradient of genetic variation along the south‐eastern U.S. coast.     

Geographic and racial variation in premature mortality in the U.S.: analyzing the disparities

Life expectancy at birth, estimated from United States period life tables, has been shown to vary systematically and widely by region and race. We use the same tables to estimate the probability of survival from birth to age 70 (S(70)), a measure of mortality more sensitive to disparities and more reliably calculated for small populations, to describe the variation and identify its sources in greater detail to assess the patterns of this variation. Examination of the unadjusted probability of S(70) for each US county with a sufficient population of whites and blacks reveals large geographic differences for each race-sex group. For example, white males born in the ten percent healthiest counties have a 77 percent probability of survival to age 70, but only a 61 percent chance if born in the ten percent least healthy counties. Similar geographical disparities face white women and blacks of each sex. Moreover, within each county, large differences in S(70) prevail between blacks and whites, on average 17 percentage points for men and 12 percentage points for women. In linear regressions for each race-sex group, nearly all of the geographic variation is accounted for by a common set of 22 socio-economic and environmental variables, selected for previously suspected impact on mortality; R(2) ranges from 0.86 for white males to 0.72 for black females. Analysis of black-white survival chances within each county reveals that the same variables account for most of the race gap in S(70) as well. When actual white male values for each explanatory variable are substituted for black in the black male prediction equation to assess the role explanatory variables play in the black-white survival difference, residual black-white differences at the county level shrink markedly to a mean of -2.4% (+/-2.4); for women the mean difference is -3.7% (+/-2.3).     

Genetic studies of low-abundance human plasma proteins. V. Evidence for a second orosomucoid structural locus (ORM2) expressed in plasma.

Orosomucoid (ORM) or alpha-1-acid glycoprotein is an acute-phase protein of human plasma whose function is suggested to be the competitive inhibition of cellular recognition by infective agents. Genetically determined variation in ORM has been reported, with two major alleles segregating in all populations studied to date. Isoelectric focusing-immunoblotting studies of ORM revealed the presence of isoprotein species that did not segregate with the predominant alleles at the ORM locus and suggested the expression of a second structural gene locus for orosomucoid (ORM2). Genetically independent variation consistent with expression of the ORM2 locus was observed in plasma samples from American blacks but was not observed in U.S. whites or sampled populations of North- and South-American Indians, Eskimos, Aleuts, or New Guinea Highlanders. The population allele frequencies for this locus were .958, .025, .006, and .011 for alleles ORM*1, ORM2*2, ORM2*3, and ORM2*4, respectively. Family studies confirm the autosomal codominant inheritance of the observed phenotypes.     

Genetic studies of low-abundance human plasma proteins. XII. A new variant of corticosteroid-binding globulin detected by isoelectric focusing/immunoblotting

Thin-layer polyacrylamide gel isoelectric focusing over the pH range 3.5-5 followed by immunoblotting was used to investigate the occurrence and frequency of genetic variation in corticosteroid-binding globulin (CBG). Plasma samples from US Caucasians (n = 105) and US Blacks (n = 106) from Pittsburgh, Pa., Canadian Indians from Vancouver Island (n = 91) and Nigerian Blacks (n = 116) were analyzed. A complex isoprotein pattern was observed in all individuals tested. Reduction of this pattern to a single primary band following neuraminidase treatment indicates that the observed intraindividual variation is due to variation in the number of sialic acid residues associated with CBG. The CBG variant pattern consisted of a series of isoprotein bands having the same mobility as the common pattern, and a second series of bands at a more acidic isoelectric point. This pattern is consistent with heterozygosity for a rare CBG allele.     

Combined biochemical and electron microscopic analyses reveal the architecture of the mammalian U2 snRNP.

The 17S U2 small nuclear ribonucleoprotein particle (snRNP) represents the active form of U2 snRNP that binds to the pre-mRNA during spliceosome assembly. This particle forms by sequential interactions of splicing factors SF3b and SF3a with the 12S U2 snRNP. We have purified SF3b and the 15S U2 snRNP, an intermediate in the assembly pathway, from HeLa cell nuclear extracts and show that SF3b consists of four subunits of 49, 130, 145, and 155 kD. Biochemical analysis indicates that both SF3b and the 12S U2 snRNP are required for the incorporation of SF3a into the 17S U2 snRNP. Nuclease protection studies demonstrate interactions of SF3b with the 5' half of U2 small nuclear RNA, whereas SF3a associates with the 3' portion of the U2 snRNP and possibly also interacts with SF3b. Electron microscopy of the 15S U2 snRNP shows that it consists of two domains in which the characteristic features of isolated SF3b and the 12S U2 snRNP are conserved. Comparison to the two-domain structure of the 17S U2 snRNP corroborates the biochemical results in that binding of SF3a contributes to an increase in size of the 12S U2 domain and possibly induces a structural change in the SF3b domain.     

Phenylketonuria in U.S. blacks: molecular analysis of the phenylalanine hydroxylase gene

We investigated the frequency, origin, and molecular basis of phenylketonuria (PKU) in U.S. blacks. On the basis of 10 years of Maryland newborn-screening data, we found the frequency to be 1/50,000, or one-third that in whites. We performed haplotype analysis of the phenylalanine hydroxylase (PAH) gene of 36 U.S. blacks, 16 from individuals with classical PKU and 20 from controls. In blacks, 20% of wild-type PAH alleles have a common Caucasian haplotype (i.e., haplotype 1), whereas 80% had a variety of haplotypes, all rare in Caucasians and Asians. One of these, haplotype 15, accounted for a large fraction (30%). Among black mutant PAH alleles, 20% have a haplotype (i.e., either haplotype 1 or haplotype 4) common in Caucasians; 40% have a haplotype rare in Caucasians and Asians, and 40% have one of two previously undescribed haplotypes. Both can be derived from known haplotypes by a single event. One of these haplotypes is characterized by a new MspI restriction site, located in intron 8, which was present in five of 16 black mutant alleles but was not present in 60 U.S. black control, 20 U.S. Caucasian control, or 20 Caucasian mutant PAH alleles. Sequence analysis of DNA from a single individual, homozygous for the new MspI associated haplotype, shows homozygosity for a C----T transition at nucleotide 896 in exon 7 of the PAH cDNA, resulting in the conversion of leucine 255 to serine (L255S).     

3H]phenamil binding protein of the renal epithelium Na+ channel. Purification, affinity labeling, and functional reconstitution

This paper describes a large-scale purification procedure of the amiloride binding component of the epithelium Na+ channel. [3H]Phenamil was used as a labeled ligand to follow the purification. The first two steps are identical with those previously described [Barbry, P., Chassande, O., Vigne, P., Frelin, C., Ellory, C., Cragoe, E. J., Jr., & Lazdunski, M. (1987) Proc. Natl. Acad. Sci. U.S.A. 84, 4836-4840]. A third step was a hydroxyapatite column. The purified material consisted of a homodimer of two 88-kDa proteins that migrated anomalously in SDS-PAGE to give an apparent Mr of 105,000. Deglycosylation by treatment with neuraminidase and endoglycosidase F or with neuraminidase and glycopeptidase F indicated that less than 5% of the mass of the native receptor was carbohydrate. Sedimentation analysis of the purified Na+ channel in H2O and D2O sucrose gradients and gel filtration experiments led to an estimated molecular weight of the [3H]phenamil receptor protein-detergent-phospholipid complex of 288,000 and of the native [3H]phenamil receptor protein of 158,000. [3H]Br-benzamil is another labeled derivative of amiloride that recognized binding sites that had the same pharmacological properties as [3H]phenamil binding sites and that copurified with them. Upon irradiation of kidney membranes, [3H]Br-benzamil incorporated specifically into a 185-kDa polypeptide chain under nonreducing electrophoretic conditions and a 105-kDa protein under reducing conditions. The same labeling pattern was observed at the different steps of the purification. Reconstitution of the purified phenamil receptor into large unilamellar vesicles was carried out. A low but significant phenamil- and amiloride-sensitive electrogenic Na+ transport was observed.